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Don't repeat the failures of the TPP in other agreements
Media Release, November 22, 2016: The failed TPP should not be used as a model for the future trade agreements, a diverse network of community organisations has warned.
The warning comes after leaders discussed future trade deals at the APEC summit in Peru this weekend, including the 16-member Regional Comprehensive Economic Partnership (RCEP)*, which includes China but not the US, and a more long-term proposal for a future Free Trade Area of the Asia Pacific (FTAAP).
AFTINET Convener Dr Patricia Ranald said the TPP's death should teach us some key lessons about future trade policy.
"Donald Trump didn't kill the TPP. His opposition was only the final blow which came at the end of more than six years of criticism from public health, environment, church, union and other community organisations," said Dr Ranald.
"These critics were not against trade itself but against unfair trade deals. Their criticism was based on human rights and environmental values.
"Even the pro-trade Productivity Commission criticised the TPP's inclusion of corporate rights to sue governments and the entrenchment of monopolies as the opposite of free trade.
"Unfortunately, leaked RCEP documents show that some governments are pushing for TPP-like provisions on stronger medicine monopolies and corporate rights to sue governments in the RCEP."
Dr Ranald said that these proposals in the RCEP are generating the same opposition as they did in the TPP.
"The death of the TPP has shown that citizens simply won't accept trade deals which entrench corporate rights and monopolies, and prevent governments from regulating in the public interest.
"These public policy issues should be debated and decided through a democratic parliamentary process, not secretly decided through trade deals.
"The failure of the TPP shows that we need more open and democratic trade processes and fairer trade agreements."
*The RCEP includes the 10 ASEAN countries plus China, India, Japan, South Korea, Australia and New Zealand
Dr Ranald's full article: Don't repeat the failures of the TPP in other agreements
AFTINET's APEC explainer: APEC: Lots of trade talk but no clear path forward


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Pink Tourmaline Bracelet
Natural stones cut into 6mm smooth rondelle stones on a hand-knotted adjustable cord bracelet.
Pink Tourmaline or Rubellite -
This is the tourmaline of love, opening one up to an awareness and experience of universal love. It helps to clear and invigorate all of our chakras. This stone brings a higher vibration providing knowledge to conquer fears and open channels between the left and right brain.
Found in Brazil, Afghanistan and Sri Lanka.

subset: commoncrawl


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GOOD Ideas for Cities: Supporting Food Truck Culture In New Orleans
Vendors and creatives have devised a plan to combat and alter the archaic laws suppressing the growth of the food truck community in New Orleans.
In recent years, New Orleans has become an urban incubator for creative ideas, drawing artists and entrepreneurs together to help guide the city towards a positive future. This meeting-of-the-minds atmosphere has revealed a simple truth about the city: New Orleans has a lot of archaic laws in dire need of a modern overhaul.
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Eneldo is a catering company with 40 years of experience, specialized in the full organization of all types of events, with constant research into new gastronomic trends, combining tradition and innovation, quality and service.
Contact person: Carlos de Miguel / Beatriz Villaescusa
Address: Avd. del Dr. Severo Ochoa, 37 - Local 4 F, Madrid . Madrid. Hours: 9:00 a 18:00 horas
Website: www.eneldo.es


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Разместите первым отзыв для этого товара
BackorderОбычно 10-20 дней
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Galaxy offers you a state of the art rescue system.
The system is designed for the rescue of crew and aircraft, specifically for 1 and 2-seat light aircraft, ultralights and experimental aircraft and lately for General Aviation aircraft and unmanned aircraft. The product is certificated by LAA ČR, USA, Australia,Canada,South Africa and the DULV, BAM Germany and complies with the conditions for purchase of rocket systems in the trade net of ČR, USA and EC. The system has earned the patent certificate PV 1859-94.
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Learn the Various Tasks Performed by Locksmiths.
Locksmiths specialize in different types of locks and keys such as door, car, windows, and safes keys and locks.These specialists also perform repair, installation, as well as adjusting the locks of cars and buildings. Also, if you are locked out, this specialist will help get you into your car or home. Locksmiths also provide consultation on security systems to their clients. At the same time, locksmiths are usually skilled to do a variety of tasks. For example, they duplicate and fabricate keys. They can also bypass locks and change lock combinations when required.
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Homes, commercial, private and government properties, as well as automobiles, need to have a good security system. The different types of locks usually give different security level. On the other hand, different types of locks will be used differently depending on the required level of security.However, you need to contact a professional locksmith South Houston to install sophisticated security systems.
usually, locksmiths save people so much time and money.With locksmith same day service, they ensure the task is performed fast thereby eliminating any delays. At the same time, a locksmith will help you get back into your car quickly especially in a location that could be dangerous.


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Kolewa Foundation entirely depends on donations.
We receive no subsidies or other forms of financing. There is always a need for money.
Fortunately, we have now built up a loyal following of regular donors. We receive donations from individuals, funds, companies and foundations.
Read more about one-off gifts, regular donations, donations for special projects.
Because we are small, we can make it very transparent where the money is going. We aim that 95% of our revenues directly benefit the projects. In our annual report, we justify ourselves every year about our plans and the spending of donations.
Kolewa has the ANBI- status which means we are a registered charity and it can be fiscally attractive (e.g. in Holland) for you to donate to Kolewa.
But there is more!
Internships and volunteer work
We can always use interns. Following training in the field of speech therapy, dietetics, physiotherapy, nursing and want to broaden your experience in a different culture?
The same goes for volunteers.
Please take a look at our page internships and volunteers
Visit us during your holiday:
Are you planning on spending your holiday in Bali? Think about a visit to Kolewa. We are happy to receive you! Read more
Kolewa cannot do without ambassadors. In the Netherlands and other countries such as Australia, we are looking for people who want to work for Kolewa. For example by helping our messages to share online, share press releases to local newspapers and – sending news. Or in your network: find individuals, corporations and foundations and make them enthusiastic for Kolewa. Or maybe you have yourself a very different good idea! Something for you? Want to know more? Contact us!


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- © 2009 American Society of Plant Biologists
In Arabidopsis (Arabidopsis thaliana), jasmonate is a key signal required for stamen and pollen maturation and thus for male fertility. Using transcriptional profiling, we have previously identified a set of 13 transcription factors that are proposed to be involved in controlling jasmonate responses in stamens. This finding suggests that a transcriptional cascade regulates the many developmental and biochemical pathways required to ensure fertility; however, the organization of this cascade is currently not understood. Here, we provide a genetic characterization of the role of MYB108 and map its relationship to MYB21 and MYB24, two other transcription factors involved in the jasmonate response in Arabidopsis stamens. Transcriptional profiling and analysis of plants expressing a MYB108:GUS fusion protein demonstrated that MYB108 expression is largely confined to sporophytic tissues of the stamen. Three allelic myb108 mutants exhibited reduced male fertility that was associated with delayed anther dehiscence, reduced pollen viability, and decreased fecundity relative to wild type. These phenotypes were all found to be exacerbated in myb108 myb24 double mutants, which also had shorter stamen filaments. Measurements of MYB108 transcript levels in wild-type and mutant flowers showed that expression of this gene is strongly dependent on MYB21. Taken together, our results indicate that MYB108 and MYB24 have overlapping functions and act downstream of MYB21 in a transcriptional cascade that mediates stamen and pollen maturation in response to jasmonate.
Flower development in angiosperms is arguably the most complex and highly regulated developmental process in the plant kingdom. It entails individual programs for the development of male and female organs (stamens and carpels, respectively) and for the haploid gametophytes that these organs produce and nurture (Goldberg et al., 1993; Ma, 2005). The accessory floral organs (sepals, petals, and pedicel) have their own developmental programs (Coen and Meyerowitz, 1991; Yanofsky, 1995). Perhaps not surprisingly, most of the major plant hormones have been proposed to have roles in floral development, including auxin (Nagpal et al., 2005; Aloni et al., 2006), gibberellins (Weiss, 2000; Yu et al., 2004), cytokinin (Lindsay et al., 2006), abscisic acid (Wang et al., 1999), ethylene (Rieu et al., 2003), and jasmonate (Feys et al., 1994; McConn and Browse, 1996). Determining the mechanisms through which these hormones act is key to understanding flower development and plant fertility.
In self-pollinating plants, such as Arabidopsis (Arabidopsis thaliana), the development and maturation of floral organs requires a high level of coordination. Anther dehiscence and the release of mature pollen must occur when the stigmas (and ovules) are receptive. In addition, elongation of stamen filaments must occur shortly before anther dehiscence to correctly position the anther immediately above the stigmatic surface for efficient pollen transfer and subsequent fertilization (Stintzi and Browse, 2000; Ishiguro et al., 2001). Mutations that disrupt steps in pollen maturation, filament elongation, or anther dehiscence often reduce male fertility or make plants completely male-sterile. Forward and reverse genetic approaches have identified many genes required for full male fertility, and functional analysis of the proteins that these genes encode has provided information about the biochemical and developmental pathways involved in stamen and pollen development (Ma, 2005).
Jasmonic acid (JA) and its chemical derivatives (collectively known as jasmonates) are oxylipin signaling molecules that are key regulators of both stress responses and development in plants. In Arabidopsis, jasmonate signaling is required for fertility, and mutants that are deficient in jasmonate synthesis (McConn and Browse, 1996; Sanders et al., 2000; Stintzi and Browse, 2000; Ishiguro et al., 2001; Park et al., 2002) or in jasmonate signaling (Feys et al., 1994; Chini et al., 2007; Thines et al., 2007) are male-sterile. These mutants show a characteristic set of developmental defects. Floral organs initially develop normally within the flower bud up to stage 12 of flower development (Smyth et al., 1990), but the anther filaments do not elongate sufficiently to place the anther locules above the stigma at the time of flower opening. The anther locules do not dehisce at the time of flower opening (although limited dehiscence does occur later) so that pollen is not released. Furthermore, although pollen on mutant plants develops to the trinucleate stage, it is predominantly inviable, with <4% of mutant pollen grains germinating (McConn and Browse, 1996; Stintzi and Browse, 2000). In jasmonate-synthesis mutants such as fad3 fad7 fad8, aos, and opr3 (but not in the jasmonate-perception mutants), application of exogenous jasmonate is able to restore fertility (McConn and Browse, 1996; Stintzi and Browse, 2000; Park et al., 2002). Restoration is very stage specific. When flower clusters were treated with a single application of jasmonate, only flower buds that were in the middle of stage 12 at the time of application produced seed; buds at earlier or later stages of development remained sterile (Stintzi and Browse, 2000). These observations suggest that jasmonate triggers developmental programs required for stamen and pollen maturation.
In the flowers, as in other parts of the plant, the action of hormones such as jasmonate is mediated by transcription factors that initiate and coordinate gene expression programs. Genetic screens have identified many transcription factors whose expression is required for flower development and function (Yanofsky, 1995; Jack, 2004; Ma, 2005). However, for most processes, including jasmonate-regulated stamen and pollen maturation, it is likely that only a subset of the relevant transcription factors have been characterized, and as a result, we have no overall understanding of the transcriptional regulation that underlies the developmental and biochemical programs involved.
One problem that has limited genetic analysis of transcription factor action is the functional overlap that is often observed between two or more transcription factors (Meissner et al., 1999; Pontier et al., 2001; Zhang, 2003). Identifying (candidate) transcription factor partners usually requires some knowledge of which transcription factors are induced (and therefore putatively active) in response to a hormone or other signal. We took advantage of our ability to trigger stamen and pollen maturation in opr3 plants with jasmonate to perform transcriptional profiling of gene expression in stamens in response to jasmonate treatment (Mandaokar et al., 2006). During the 22 h of the experiment, a total of 146 genes encoding known or putative transcription factors were induced or repressed in jasmonate-treated stamens compared with controls. A series of considerations and analyses refined this extended list to a set of 13 genes that we conclude are most likely to be key regulators of the stamen and pollen maturation processes triggered by jasmonate. These genes are strongly induced in opr3 stamens following jasmonate treatment and are highly expressed in wild-type stamens relative to stamens from untreated (sterile) opr3 flowers. Most of the 13 transcription factors identified appear to act specifically in flower development. For example, expression of the MYB21 gene in stamens was induced nearly 400-fold by jasmonate and myb21-1 null mutants showed similar symptoms of male sterility as described for opr3 and several other jasmonate mutants (Mandaokar et al., 2006). Interestingly, null mutations in the MYB24 gene, which is closely related to MYB21, do not lead to any detectable reduction in fertility or other difference from wild type, although in the myb21-1 background myb24 mutations did exacerbate the effects on flower development and fertility (Mandaokar et al., 2006). Mutant analyses of two additional genes from our list have independently identified MYB32 and IAA19 as having roles in stamen or pollen development, because mutations in either of these genes results in reduced male fertility (Preston et al., 2004; Tatematsu et al., 2004).
Among the transcription factors identified in our profiling experiments, MYB108 (At3g06490) was an additional promising candidate. MYB108 transcript was increased more than 60-fold (relative to the zero-time control) at 22 h after jasmonate application. Treatment of flowers with 12-oxo-phytodienoic acid (OPDA), a precursor of jasmonate that does not restore fertility to opr3 plants, did not result in a significant increase of MYB108 transcript. Finally, transcript levels of MYB108 in wild-type stamens were 20-fold higher than those in opr3 stamens that were either untreated or treated with OPDA (Mandaokar et al., 2006). Here, we report that MYB108 is a JA-inducible transcription factor gene with an important role in stamen development and male fertility in Arabidopsis. Reverse-genetic analysis demonstrates that MYB108 is required for correct timing of anther dehiscence. In combination with MYB24, it regulates three aspects of male fertility: filament elongation, anther dehiscence, and pollen viability. Results from additional genetic analysis indicate that MYB108 expression is regulated by the upstream transcription factor, MYB21. We propose that MYB108 and MYB24 are two critical components of the JA-mediated transcriptional cascade that regulates final stages of stamen development and male fertility.
Identification of myb108 T-DNA Insertion Mutants
Phylogenic analysis of the R2R3-MYB proteins in Arabidopsis places MYB108 in subgroup 20 of this transcription factor family (Stracke et al., 2001). Subgroup 20 has five other members, MYB78, MYB112, MYB2, MYB62, and MYB116 (Fig. 1A ). The MYB21 and MYB24 transcription factors that have demonstrated roles in jasmonate regulation of stamen and pollen maturation (Mandaokar et al., 2006) are the two members in subgroup 19 and thus closely related to the subgroup 20 proteins.
To examine the possible role of MYB108 in stamen development and fertility, we used a reverse genetic approach to identify homozygous T-DNA and transposon insertion mutants. We identified three insertion lines listed for MYB108 in The Arabidopsis Information Resource database (www.arabidopsis.org): SALK_056061, SALK_024059, and CSHL_GT6213 (Martienssen, 1998; Alonso et al., 2003). These three lines were obtained from the Arabidopsis Stock Center at The Ohio State University and characterized as described under "Materials and Methods." Seed was collected from one homozygous plant of each line and designated myb108-1 (SALK_056061), myb108-2 (SALK_024059), and myb108-3 (CSHL_GT6213). myb108-1 and myb108-2 are in the Columbia (Col-0) background, while myb108-3 is derived from the Landsberg erecta (Ler) ecotype. RNA prepared from developing flower buds of mutant plants was examined by reverse transcription (RT)-PCR using the MYB108F and MYB108R primers together with primers to the ACTIN2 gene as a control. We could not detect the presence of any full-length transcript for MYB108 in any of the three mutant lines (Fig. 1B), even though a strong PCR band was observed using RNA from wild-type flowers and ACTIN2 transcript was detected in all the RNA samples.
The three T-DNA insertions are all in the first intron of the MYB108 gene (Fig. 1B) and are predicted to preclude translation of both the R2 and R3 domains. Thus, it is highly unlikely that any truncated mRNA transcribed from the mutant gene would retain any biological function. T-DNA inserts in intronic sequences are sometimes spliced out of the pre-mRNA with the intron, thus allowing for some gene expression. However, our inability to detect MYB108 transcript in the three mutant alleles indicates that each of the mutations is null.
When plants of all three mutant lines were grown side-by-side with wild-type controls, we could not detect any phenotype during vegetative growth. However, careful observation of newly opened flowers revealed a consistent phenotype for all three myb108 alleles. Although the stamen filaments of myb108 flowers had elongated to place the anthers level with or slightly above the stigmatic surface at the time of flower opening, anther dehiscence did not occur but instead was delayed by approximately 12 h relative to the corresponding Col-0 or Ler wild type. This defect in anther dehiscence was associated with delayed senescence of stamens, petals, and sepals of flowers and reduced seed set for all three myb108 alleles. Delayed senescence is often observed in male-sterile lines. The phenotype and seed set for myb108-1 are shown in Figure 1, C to E. The delayed senescence of flower parts is reflected by the larger number of open flowers on myb108 compared with wild type (Fig. 1C), while the smaller siliques on myb108 (Fig. 1, C and D) indicate reduced seed set. Seed counts from 80 randomly selected siliques confirmed the reduced seed set with myb108 siliques containing 32.2 ± 1.4 (mean ± sem) seeds compared with 54.0 ± 0.8 for wild-type controls (Fig. 1E).
Expression Pattern of the MYB108 Gene
Our previous results (Mandaokar et al., 2006) and data on MYB108 expression from the Arabidopsis Gene Expression Atlas (Schmid et al., 2005) indicates that this gene is flower specific and is predominantly induced in stamens at stages 12 and 15 of flower development. To confirm and extend these results, we first used quantitative, real-time PCR (qPCR) to reassess the induction of MYB108 transcript in stamens following treatment of opr3 flower buds with jasmonate or OPDA. In this experiment, MYB108 was induced approximately 100-fold at 22 h after jasmonate treatment, but there was no response to OPDA (Fig. 2A ). To determine if MYB108 expression is dependent on activity of the SCFCOI1 ubiquitination complex, we used RNA gel-blot analysis to measure MYB108 transcripts in flowers of wild-type and coi1-mutant flowers that had been treated with 100 μm jasmonate or with a control solution. The results show high expression of MYB108 in wild-type flowers with or without jasmonate treatment. By contrast, the MYB108 transcript was not detected in coi1 flowers even after jasmonate treatment (Fig. 2B). Taken together, these results indicate that MBY108 is induced in stamens of wild-type flowers to levels comparable to those found in opr3 stamens treated with jasmonate and that this induction is COI1 dependent.
The myb108-3 allele (derived from CSHL_GT6213) was generated in the Ler genotype by transformation with a gene-trap construct based on the maize Dissociation (Ds) transposon (Martienssen, 1998). The construct is designed to generate in-frame protein-GUS fusions when the transposon inserts into an intron. Thus, heterozygous myb108-3 plants are phenotypically wild type and allow analysis of the tissue specificity of MYB108 expression. We performed histochemical staining for GUS activity at various stages of development from germination to maturity. GUS staining was observed very specifically in the stamens of flowers, particularly in the anthers (Fig. 2C). The intensity of staining increased progressively during flower maturation and opening (Fig. 2D), with maximum expression at stage 15 of flower development. Cross sections of anthers showed GUS staining throughout the maternal tissues with strongest expression in the vascular and connective tissue where the anther attaches to the filament (Fig. 2E). No GUS staining was observed in any other flower or vegetative organs, except for limited staining in the axial region of seedling leaves (data not shown). The results from these GUS reporter plants are consistent with data on the expression of MYB108 transcript, described above, and indicate that MYB108 is likely specifically involved in regulation of late stages in stamen development.
myb78 and myb112 Mutations Do Not Enhance the myb108 Phenotype
The reduced fertility of myb108 mutants, the strong induction of MYB108 expression in response to jasmonate treatment of opr3 stamens, and the anther-specific expression of the myb108-3:GUS reporter all indicate that the MYB108 transcription factor contributes to the regulation of stamen maturation and male fertility in response to jasmonate signaling. However, the reduction in fertility observed in myb108 mutants is much less severe than in myb21, opr3, and other jasmonate mutants (Mandaokar et al., 2006). We therefore considered other MYB transcription factors that might have overlapping function(s) with MYB108.
All the proteins in subgroup 20 of the R2R3-MYB family contain a diagnostic WXPRL sequence in addition to the R2 and R3 domains and show higher sequence similarity to each other than to MYB proteins in other subgroups of the R2R3-MYB family (Stracke et al., 2001). The MYB2 protein has been shown to be a transcriptional activator of the dehydration-responsive gene rd22 (Abe et al., 2003) and to act in abscisic acid signaling. It is therefore unlikely to have a function that overlaps with the function of MYB108. MYB116 and MYB62 are the subgroup 20 proteins most distantly related to MYB108 with sequence identities of 47% and 43%, respectively, and the corresponding genes are not strongly expressed in stamens of developing flower buds (data not shown). We considered these unlikely candidates for a shared role with MYB108 and did not investigate them further. MYB78 (At5g49620) is the closest homolog of MYB108 (Fig. 1A) with 66% sequence identity, although data from the Arabidopsis Gene Expression Atlas (Schmid et al., 2005) indicate that this gene is expressed at only very low levels throughout plant development. On the other hand, MYB112 (At1g48000) shows 62% amino acid identity to MYB108 and is expressed in stamens and other organs of stage 15 flowers.
We obtained T-DNA insertion mutants of MYB78 (SALK_085369) and MYB112 (SAIL_562_F10) and identified homozygous lines by PCR genotyping using the same strategy described above for myb108 (Fig. 3, A and B ). RT-PCR analysis using RNA from wild type, myb78, and myb112 showed that each mutant lacked a full-length transcript of the corresponding gene (Fig. 3, A and B). Both the myb78 and myb112 knockout mutants were normal in growth, flower development, and seed set. We crossed each mutant with myb108 plants and derived homozygous myb108 myb78 and myb108 myb112 double mutant lines. When these mutants were grown together with myb108, we did not detect differences in anther dehiscence or seed set between the double mutants and the parental myb108 plants. These results indicate that it is unlikely that MYB78 or MYB112 has a function similar to MYB108.
Mutations in myb24 Enhance Sterility of myb108 Plants
We have not investigated the MYB transcription factors from subgroup 20 of the R2R3 family any further, because the low expression of these genes in stamens of wild-type flowers and especially the lack of significant induction by jasmonate in our transcriptional profiling experiment suggest that they are not involved in regulation of jasmonate responses. Instead, we next considered the possibility that the regulatory function of MYB24 overlaps with that of MYB108. The genes encoding these two transcription factors are very highly induced by jasmonate treatment of opr3 flowers (Fig. 2A; Mandaokar et al., 2006). Furthermore, the myb24-1 null mutant is fully fertile and indistinguishable from wild type in growth and development (Mandaokar et al., 2006), indicating that MYB24 function may be fulfilled by one or more other transcription factors in myb24 mutants.
Four insertion alleles of myb24 were identified in our previous work, but only myb24-1 (isolated through the Arabidopsis Knockout Facility at the University of Wisconsin) was characterized in detail. We therefore prepared RNA samples from flower buds of myb24-1, myb24-2 (homozygous derivative from SALK_030452), and myb24-3 (homozygous derivative from SAIL_284_F01) and used them as templates for RT-PCR using primers MYB24F and MYB24R designed to the 5′ and 3′ ends, respectively, of the MYB24 coding sequence. As shown in Figure 3C, these primers detected full-length MYB24 transcript in RNA from wild-type controls but not in any of the three myb24 alleles. We conclude that each of these mutant lines contains a null mutation in the MYB24 gene.
A cross between myb108-1 and myb24-1 produced F1 plants that were allowed to produce selfed F2 seed. A total of 62 F2 plants were analyzed and four of these showed greatly reduced seed set compared with myb108 and wild type. The 62 plants were genotyped by PCR (using primer combinations that distinguish wild-type and mutant alleles of the two genes) and only the four plants with very low seed set were homozygous myb108 myb24 double mutants. The 58:4 segregation in this experiment is a good fit to the 15:1 ratio (χ2 = 2.7; P > 0.1) expected for two recessive loci. We also made double mutants between myb108-2 and myb24-2 and between myb108-2 and myb24-3. In both these cases, the double mutant segregants in the F2 population had greatly reduced seed set relative to the myb108 parental allele. The consistency of this result indicates that the reduced fertility of the myb108 myb24 mutants is the result of mutations in only these two genes and is not affected by any other mutations that might potentially be present in the individual myb108 or myb24 lines. The detailed characterization reported below uses results from the myb108-2 myb24-2 mutant, and consistent results were obtained from the two additional lines.
The cumulative effect of the myb108 and myb24 mutations was evident in the phenotype of flowering shoots and sample siliques from myb108 myb24 plants (Fig. 1, C and D). Seed counts from 80 randomly selected siliques of myb108 myb24 indicated that seed production was only 13.1 ± 0.6 (mean ± sem) seeds per silique compared with 32.2 ± 1.4 for the myb108 single mutant (Fig. 1E). When myb108 myb24 flowers were pollinated with wild-type pollen, seed set was normal. However, when pollen from double-mutant plants was used to pollinate emasculated, wild-type flowers, seed set was consistently very low (Table I ). These results indicate the myb108 myb24 plants are female-fertile but male-sterile and are consistent with the notion that the mutations substantially block jasmonate responses in stamens of double-mutant plants.
We treated myb108, myb108 myb24, and opr3 flower buds with jasmonate by dipping the apical bud clusters in a solution of 100 μm methyl jasmonate. Following this treatment, opr3 plants produced three to six elongated siliques filled with seed, as expected for this mutant (Mandaokar et al., 2006). By contrast, jasmonate treatment did not enhance seed set on myb108 or myb108 myb24 plants. This lack of response is consistent with the mutations in MYB108 and MYB24 blocking jasmonate actions downstream of the initial responses mediated by SCFCOI1 and the JAZ proteins (Chini et al., 2007; Thines et al., 2007).
Delayed Anther Dehiscence in myb108 myb24 Double Mutants
Because we had observed delayed anther dehiscence in the myb108 mutant, we speculated that the myb24 mutations might further delay dehiscence and set out to investigate this process in detail. As in other jasmonate mutants (Feys et al., 1994; McConn and Browse, 1996; Sanders et al., 2000; Ishiguro et al., 2001), the timing of flower development and anthesis in myb108 and myb108 myb24 mutants is indistinguishable from wild type. Morphological changes are limited to the stamens and to delayed dehiscence of the petals and sepals, which is a result of reduced fertilization (Butenko et al., 2003). We grew wild-type, myb108, and myb108 myb24 plants together and measured the timing of anther dehiscence in the flowers. The most mature unopened flower bud in each flower cluster was identified (flower 0), and the set of six open flowers below it was treated as a developmental series (Fig. 4A ). The unopened flower buds contained no dehisced anthers, but the earliest open flowers on wild-type plants consistently had six dehisced anthers (Fig. 4B). By contrast, in myb108, anther dehiscence did not occur in the first and second open flowers. A more pronounced delay was observed in the myb108 myb24 double mutants, with no anther dehiscence occurring until flower 4 and with full dehiscence in flower 5, which corresponds to a stage when pistil elongation has already begun.
Reduced Filament Elongation of myb108 myb24 Stamens
A second important aspect of the myb108 myb24 phenotype was discovered through measuring stamen lengths. In flowers 1 through 4 in the series, stamen length for myb108 myb24 was slightly but significantly less than for wild type, with myb108 indistinguishable from wild type (Fig. 5A ). Because stamen length is particularly important to position anthers for releasing the pollen onto the stigmatic surface, we also measured carpel lengths and calculated the stamen:carpel length ratio. The results in Figure 5B show that in wild type, this is greater than one in all the open flowers, while the ratio for myb108 myb24 is consistently less than one.
The combined effects of delayed anther dehiscence and reduced stamen elongation mean that pollen in myb108 myb24 flowers is inefficiently transferred to the stigmatic surface and is often deposited on the side of the style (Fig. 5C). Microscopic examination of myb108 myb24 flowers typically revealed 20 to 50 (occasionally up to 100) pollen grains on the stigmatic surface compared with several hundred pollen grains on the stigmas of wild-type flowers. Taken together, these results show that stamens on the double mutant are inefficient at transferring pollen to the stigma.
Pollen from myb108 myb24 Mutants Has Reduced Viability and Germination
The poor seed set achieved using myb108 myb24 as a pollen donor (Table I) implies that myb108 myb24 pollen is defective. To investigate this more closely, we examined pollen viability using double staining with fluorescein diacetate and propidium iodide. Fluoroscein diacetate is taken up by living cells and converted to fluoroscein, which emits blue-green light under UV irradiation (Heslop-Harrison and Heslop-Harrison, 1970). Propidium iodide is excluded from living cells but labels dead cells with red-orange fluorescence under UV irradiation (Regan and Moffatt, 1990). The results in Figure 6 show the greatly reduced viability of myb108 myb24 pollen relative to pollen from either wild-type or myb108 flowers. Counts of fluoroscein-stained pollen grains from 10 sample microscope fields in each of three separate experiments indicated that viability of myb108 myb24 pollen by the criterion of this vital stain was consistently 20% to 25% while wild-type pollen was ≥90% viable.
To further test pollen viability and function, we measured pollen germination on artificial media (Pickert, 1988). Pollen from the mutants was transferred to the agar media either from newly dehiscent anthers (flowers 4 and 5 in Fig. 4) or by cutting open anthers from full open flowers (flowers 2 and 3). In either case, germination of myb108 myb24 was considerably lower than germination of wild-type or myb108 pollen (Fig. 6B). For wild type, counts from 10 samples (>1,000 pollen grains in total) showed an average of 80% germination. By contrast, <10% germinated pollen grains were observed in 20 samples (>1,000 sample grains) from myb108 myb24 stamens.
MYB108 Expression Is Dependent on MYB21
Available evidence indicates that MYB21 is a key regulator of the transcriptional cascade that brings about jasmonate-dependent maturation of stamens and pollen in Arabidopsis, and myb21 mutant plants have severely reduced fertility (Mandaokar et al., 2006). Following jasmonate treatment of opr3 stamens, the MYB21 gene is induced earlier than either MYB24 or MYB108, suggesting that the MYB21 transcription factor may control the expression of one or both of these genes.
As part of an investigation of MYB21 function, we conducted transcriptional profiling of genes expressed in stamens of myb21 mutant plants after treatment with 100 μm jasmonate. Comparison of gene expression in myb21 with expression in wild-type, opr3 plants treated with jasmonate, and opr3 plants untreated or treated with the inactive precursor, OPDA, will allow us to evaluate the extent to which MYB21 controls the jasmonate response in stamens. Using the data from Affymetrix arrays as a digital northern (Toufighi et al., 2005), it is clear that both MYB108 and MYB24 are highly expressed in wild-type stamens, approximately 700 expression units when the arrays are normalized to a median of 125 (Mandaokar et al., 2006; Fig. 7A ). However, in stamens from myb21, MYB108 expression is only 112, indicating that expression of this gene is very dependent on the MYB21 transcription factor. Expression of MYB24 is also decreased in the myb21 mutant but to a much smaller extent, indicating that MYB24 expression is substantially independent of MYB21. Comparisons with untreated opr3 stamens indicates that MYB24 is induced 140-fold in the myb21 mutant background, while MYB108 is induced <4-fold.
We carried out a second investigation of MYB108 expression using qPCR to measure MBY108 transcript levels in stamens of wild-type and mutant plants (Fig. 7B). The results confirm that MYB108 expression is greatly reduced in the myb21 mutant relative to wild type. MYB108 expression is decreased only to a small extent by the myb24 mutation, but, in the myb21 myb24 double mutant and in the opr3, mutant MYB108 transcripts are lower than in myb21 and only 10% to 15% of those in wild type. Taken together, these results indicate that MYB24 induction by jasmonate is largely independent of MYB21 but that MYB21 and MYB24 both contribute to induction of the MYB108 gene.
The role of jasmonate in plant reproduction has been established through the identification and characterization of mutants deficient in jasmonate synthesis or perception (Feys et al., 1994; McConn and Browse, 1996; Sanders et al., 2000; Stintzi and Browse, 2000; Ishiguro et al., 2001; Park et al., 2002; von Mallek et al., 2002). Our previous transcript profiling experiments showed that jasmonate initiates a transcriptional cascade in Arabidopsis stamens that is required for stamen maturation and male fertility (Mandaokar et al., 2003, 2006). Using reverse genetics, we have previously established a role for two early transcription factors MYB21 and MYB24 in this cascade. Mutation of MYB21 severely reduces fertility in Arabidopsis in that anther filaments are short, anthers do not dehiscence, and pollen viability is also affected to a large extent. myb24 mutants are phenotypically wild type, but this mutation exacerbates the phenotype of myb21 (Mandaokar et al., 2006). Here, we show that another MYB transcription factor, MYB108, plays an overlapping role with MYB24 to regulate late stages of stamen development and male fertility.
In our search for additional transcription factors in the jasmonate signaling cascade, we identified MYB108 as an important candidate gene, because this gene is very strongly induced between 8 and 22 h after jasmonate treatment of opr3 stamens, it is not detectably expressed in untreated or OPDA-treated opr3 plants (Mandaokar et al., 2006), and it is highly expressed in wild-type stamens. Reverse-genetic analysis using T-DNA insertion mutants showed that MYB108 is required for correct timing of anther dehiscence (Fig. 4B), a critical part of anther development, and that loss of MYB108 results in reduced fertility.
Our examination of GUS staining in the myb108-3 gene-trap mutant revealed that MYB108 expression is tightly constrained to the maternal tissue of the stamen and to a short period of time corresponding to stages 12 to 15 of flower development. No GUS staining of pollen was observed, nor of any other floral organs. The primary phenotype of myb108 is delayed dehiscence and in Arabidopsis dehiscence is believed to be facilitated by water movement out of the anther via the vasculature (Bonner and Dickinson, 1990; Ishiguro et al., 2001). The expression of MYB108 in the connective tissue and vascular bundles of the anther is thus consistent with its regulating genes required for water movement and anther dehydration.
Expression data from the Arabidopsis Gene Expression Atlas (Schmid et al., 2005) indicates that MYB108 transcript is modestly induced by salt treatment and during osmotic stress. Interestingly, a specific mutation at the MYB108 locus, botrytis susceptible1 (bos1), was isolated on the basis of its susceptibility to Botrytis cinerea, and in wild-type plants, MYB108/BOS1 is induced in Botrytis-infected tissue (Mengiste et al., 2003). As well as being susceptible to some pathogens, bos1 plants were shown to be impaired in tolerance of water deficit, salt, and oxidative stress. However, bos1 plants do not have reduced fertility and do not show delayed anther dehiscence (T. Mengiste, personal communication). We are currently unable to fully explain the differences in reproductive phenotype between our three myb108 null alleles and the bos1 mutant. However, the bos1 mutant is unusual in that homozygous and heterozygous mutant plants express very high levels of apparently full-length MYB108 transcript in vegetative tissue (Mengiste et al., 2003). The T-DNA in bos1 is inserted immediately 5′ to the ATG start codon of the open reading frame and there is also a 314-bp deletion that involves the 151-bp 5′ untranslated region and 163 bp of the promoter (Mengiste et al., 2003). A reasonable explanation for the full fertility of bos1 plants would be the production of at least some full-length, functional MYB108 protein in bos1 anthers.
Considering the modest reduction in fertility of the myb108 null mutants, we considered the possibility that additional transcription factors might also be involved. To test this possibility, we made double mutants with the closely related homologs of MYB108. Null mutants for two closely related transcription factors, myb78 and myb112, showed wild-type phenotypes. When we generated double mutants with myb108, both myb108 myb78 and myb108 myb112 failed to show any exacerbation of the myb108 phenotype, suggesting that these genes are not involved in jasmonate-regulated anther development processes. Consistent with these results, our transcript profiling experiments showed no induction for either of these genes by jasmonate (Mandaokar et al., 2006). However, the generation of double mutants between myb108 and myb24 demonstrated the related contributions that the MYB108 and MYB24 transcription factors make in controlling the jasmonate response in stamens, because all the myb108 myb24 lines showed greatly reduced fertility relative to either of the parental mutants.
All three aspects of jasmonate-regulated stamen function are more strongly affected in the myb108 myb24 double mutants than in the myb108 parental lines. Filament elongation is not significantly affected in myb108, but in the double mutant it is reduced sufficiently to compromise positioning of the anthers above the stigma. A substantial delay in anther dehiscence is the most obvious aspect of the myb108 phenotype (Fig. 4B), and the delay is longer in myb108 myb24 flowers. Pollen germination is reduced by approximately 35% in myb108 relative to wild type, but by 90% in the double mutant (Fig. 6). These results indicate that MYB108 and MYB24 both function to activate jasmonate-responsive genes required for correct stamen and pollen maturation.
Because Arabidopsis flowers produce pollen in considerable excess (Jürgens et al., 2001), the severely deficient pollen and stamen functions in myb108 myb24 still allow some seed set (Fig. 1, C–E). By contrast, mutants that are completely blocked in jasmonate synthesis or jasmonate signaling, such as opr3, coi1, myb21, and the JAZ1Δ3A transgenic plants, are more completely sterile (Feys et al., 1994; Stintzi and Browse, 2000; Mandaokar et al., 2006; Thines et al., 2007). We infer that one or more additional transcription factors may partially compensate for the loss of MYB108 and MYB24 function.
We have recently proposed that the active form of jasmonate is jasmonoyl-Ile, which acts by enhancing the affinity of SCFCOI1 ubiquitin ligase for JAZ repressor proteins (Thines et al., 2007). This suggests a model in which ubiquitination and subsequent degradation of JAZ proteins leads to activation of primary transcription factors that then induce the expression of additional transcription factors and other components of the jasmonate-induced pathways. We propose that, in flower organs, the MYB21 and MYB24 genes are principal targets of this signaling. The results reported here, together with our previous results (Mandaokar et al., 2006; Thines et al., 2007), indicate a complex relationship among the transcription factor genes and proteins that mediate jasmonate responses in stamens. The data in Figure 7 indicate that MYB21 is one of the transcription factors contributing to maximum expression of the MYB24 gene. MYB21 or MYB24 is required for expression of MYB108, and both of these transcription factors are necessary for maximum accumulation of MYB108 transcript. The timing of gene induction following jasmonate treatment of opr3 stamens is consistent with these observed dependencies. Transcriptional profiling indicates that MYB21 is induced first, followed by MYB24 and finally MYB108 (Mandaokar et al., 2006; Fig. 2A). Additional MYB transcription factors are induced by jasmonate and, at least in some cases, required for full male fertility (Preston et al., 2004; Tatematsu et al., 2004; Mandaokar et al., 2006). In plants and other organisms, the MYB transcription factors have often been shown to regulate gene transcription in concert with bHLH proteins, which include the MYC transcription factors. Our transcript profiling experiments identified early- and late-induced bHLH genes, including those encoding bHLH31 (At1g59640), bHLH152 (Atlg22380), and bHLH145 (At5g50010; Mandaokar et al., 2006). It is possible that regulation of the expression of these genes involves interactions analogous to those described here for the MYB genes. Indeed, the interactions may extend between the MYB and bHLH classes and to other transcription factors involved in the jasmonate response in stamens. One explanation for such a complex interrelationship is that it allows robust regulation of stamen and pollen maturation processes that are essential for reproductive fitness.
Plant Material and Methyl Jasmonate Treatment
The T-DNA insertion lines in the Col-0 ecotype of Arabidopsis (Arabidopsis thaliana) Hegn., SALK_056061, SALK_024029, SALK_030452, SALK_01722, and SAIL_284_F10, were identified using the SIGnAL database (http://signal.salk.edu/cgi-bin/tdnaexpress) and obtained from the Arabidopsis Biological Resource Center at The Ohio State University. The gene trap line CSHL_6213 was obtained from Cold Spring Harbor Laboratory and is in the genetic background of the Ler ecotype. Wild-type and T-DNA insertion mutant seeds were sown on soil and stratified at 4°C for 2 d. The plants were gown in a controlled growth chamber in a continuous light condition (150 μmol m−2 s−1) at 22°C. The opr3, myb108, and myb108 myb24 mutants were treated by spraying separately with 0.1% Tween 20 (control) and 0.03% methyl jasmonate (Bedoukian Research) in 0.1% Tween 20 on inflorescences. Development of siliques was observed 1 week after the treatment.
Isolation of Homozygous T-DNA Insertion Mutants
Screening of homozygous insertion mutants was done by genomic PCR using a combination of gene specific and T-DNA left border (LB) primers. Approximately 12 plants of each line were grown on soil. Using DNA from each plant and oligonucleotide primers designed to the 5′ and 3′ ends of the MYB108 coding sequence (MYB108F and MYB108R, respectively) and primers matching sequences in the LB of the T-DNAs or in the transposon, we confirmed the site of insertion in each line by PCR and identified plants that were homozygous for each mutation. LB primer for SALK lines, 5′-TGGTTCACGTAGTGGGCCATCG-3′; for line GT6213, 5′-ACCCGACCGGATCGTATCGGT-3′; and for SAIL lines is LB3, 5-TAGCATCTGAATTTCATAACCAATCTCGATACAC-3′. Gene-specific forward and reverse primers for MYB108: MYB108F 5′-AATGGAGAAGGTCGCTGGAACTCT-3′ and MYB108R 5′-AACCAGCTGAGGTTACTCTGCTCT-3′; MYB78: MYB78 F 5′-ATGGGTGACAAAGGAAGGAGCTTAA-3′ and MYB78R, 5′-TCAGAAGCTTCCATTGTCGTGGAC-3′; MYB112: MYB112F 5′-AGAAGAAGTCGAAGAAGTCGAGAA-3′ and MYB112R 5′-CTACTGTATGAGCCACTTGTTGAGC-3′; and MYB24: MYB24F 5′AGAGAAAGTAGTGGTGGGTCTGGA3′ and MYB24R 5′-GCCAAAGATCATCGACGCTCCAAT-3′.
Promoter-GUS Fusion and Histochemical Analysis
The gene trap line from CSHL was used to study the staining pattern of GUS in various tissues. These lines were developed by transposon mutagenesis in Arabidopsis using Ds from maize (Zea mays) that has been engineered to carry a uidA (GUS) reporter gene and an NPTII kanamycin resistance gene. In gene trap construct, the reporter gene is preceded by a triple splice acceptor and by a short intron so that insertion into chromosomal introns leads to reporter gene expression via alternate splicing in each reading frame (Martienssen, 1998). Following transposition, GUS will express if it is integrated in the gene. The promoter of the genes drives the expression of GUS. Gene trap line GT6213 is obtained from the Cold Spring Harbor Laboratory. The insertion is in the first intron of the MYB108 gene. Because the reporter carries a triple splice acceptor, its insertion will result in a translational fusion with the first exon of MYB108. The MYB108 promoter then regulates the expression of this translation fusion.
The staining of GUS was done according to standard protocol (Weigel and Glazebrook, 2002). Individual flowers from plants were removed and placed in the GUS substrate buffer (50 mm sodium phosphate buffer, pH 7.0, 0.1% Triton X-100, 3 mm potassium ferricyanide, 3 mm potassium ferrocyanide, and 1 mm 5-bromo-4-chloro-3-indolyl-β-glucuronic acid). After a 5-min vacuum infiltration, samples were placed in a 37°C incubator overnight. The next day, tissue was fixed in 50% ethanol, 5% acetic acid, and 3.7% formaldehyde for overnight and dehydrated through an ethanol series. Samples were processed through ethanol to CitriSolv and then into paraffin. The paraffin-embedded samples sat at 62°C for 3 d, after which blocks were cast. Longitudinal and transverse sections of anthers (15 μm) were deparaffinized in CitriSolv, dehydrated in oil under a coverslip, and observed under a microscope.
RT-PCR and qRT-PCR
Total RNA from flowers of wild type and all the mutants was isolated using Trizol and purified using an RNeasy kit (Qiagen) according to the manufacturer's instructions. Transcript presence in wild-type, myb108, myb78, myb112, and myb24 plants was measured by one-step RT-PCR (Invitrogen) with 100 ng of RNA isolated from flowers using the same set of primers used for genomic PCR. The PCR conditions were 42°C for 50 min and 30 cycles of the following: 94°C for 20 s, 58°C for 30 s, and 72°C for 2 min (the final synthesis step was extended for 5 min). Amplification products were analyzed with agarose gel electrophoresis.
For qRT PCR, total RNA was isolated from inflorescence of wild-type, opr3, myb21-1, myb24-1, and myb21-1 myb24-1 plants, and first-strand cDNA synthesis was performed on 2 μg of total RNA using SuperScriptIII cDNA synthesis kit (Invitrogen) according to the manufacturer's instructions. Reaction was performed in a 20-μL volume and diluted 2-fold after the completion of reaction. qRT-PCR was performed on a Stratagene MX3000p using the SYBR Green I dye method (Stratagene). Reaction mixture (20 μL) contained 2 μL of the first-strand cDNA, 2.0 μL 1× PCR buffer, 1.5 mm MgCl2, 0.2 mm dNTPs, 0.2 μm of each sequence-specific primers, 0.6 μL of ROX dye (diluted to 1:500), and 10 μL SYBR Green mix (Invitrogen) with Taq polymerase. The amplification protocol was 95°C (2 min) and 40 cycles of amplification cycle (95°C [15 s], 55°C [30 s], and 72°C [30 s]). All the experiments were carried out in triplicate and repeated twice on two biological samples. MYB108 mRNA level was determined by normalizing with the ACTIN2 cDNA of each sample. The relative transcript level of MYB108 in opr3, myb21-1, myb24-1, and myb21-1 myb24-1 line was compared using wild-type (Col-0) as a calibrator. The gene-specific primer sequences used for MYB108 were forward 5′-AATGGAGAAGGTCGCTGGAACTCT-3′ and reverse 5′-CGTTGTCCGTTCTTCCCGGTAAAT-3′; the primer sequences for ACTIN2 were forward, 5′-GGTGATGGTGTGTCTCACACTG-3′ and reverse, 5′-GAGGTTTCCATCTCCTGCTCGTAG-3′. The quality control was carried out using both electrophoresis analyses on a 1.5% agarose gel and dissociation curve analysis performed immediately after the end of amplification.
RNA-Blot Analysis
Flowers of wild-type and coi1 plants were treated with 100 μm methyl jasmonate in 0.1% Tween 20 solution. Control treatment contained only 0.1% Tween 20. Samples were collected 24 h after treatment. RNA extraction and RNA gel-blot analysis were performed as described previously (Mandaokar et al., 2003).
Pollen Viability and Germination Test
Double staining with fluorescein diacetate and propidium iodide was performed using the method of Regan and Moffatt (1990). A stock solution of 2 mg /mL fluorescein diacetate was made in acetone and added drop wise to 17% Suc (w/v) until the solution became cloudy. Propidium idodite was diluted to 1 mg/mL in water and diluted to 100 μL/mL with 17% Suc (w/v). Equal amounts of fluorescein diacetate and propidium iodide solutions were mixed together and were added to freshly isolated pollen on a glass slide. The pollen were covered with a coverslip and viewed under UV light under a microscope (Olympus IX70).
Pollen germination assay was done according to the methods of Pickert (1988). Pollen was isolated from mature flowers by gently releasing them from the anther locules onto a petri dish (5 cm) containing pollen germination medium. Pollen from myb108 and myb108 myb24 mutants was released by manually opening the anther locule. The medium consisted of tap water, 20% (w/v) Suc, 80 μg mL−1 boric acid, and 10 μg mL−1 myoinositol at pH 5.8 to 6.0. The plates were incubated in the dark for 5 to 10 h at room temperature, and pollen germination was observed in a microscope (Leica MZ8).
Measurements of Stamen Length
To measure the stamen and carpel length of wild type, myb108, and myb108 myb24, sepal and petals were removed from unopened flower buds and open flowers. The series of pictures of stamen and carpel were taken under the microscope at a fixed magnification. From these images, the length of stamens and carpel was measured. The actual length of stamen was calculated by dividing the magnification.
We thank Craig Whitney for taking care of plants and Jim Wallis for helpful comments on this manuscript.
The author responsible for the distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) is: John Browse (jab{at}wsu.edu).
↵1 This work was supported by the U.S. Department of Energy (grant no. DE–FG02–99ER20323) and by the Agricultural Research Center at Washington State University.
↵2 Present address: DuPont Knowledge Centre, ICICI Knowledge Park, Turkapally, Shamirpeth Mandal, Hyderabad, India 500 078.
↵[OA] Open access articles can be viewed online without a subscription.
- Received November 14, 2008.
- Accepted December 3, 2008.
- Published December 17, 2008.


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Pinot's Palette brings art to the masses in Fairborn, Dayton Mall area
Until a pivotal moment, Tonya Shine didn't believe she had an ounce of artistic talent. Straight lines and stick figures, yes. Vividly colorful paintings worthy of hanging on walls, no way. That changed when, as a birthday gift that October evening, her friends threw her a party at Pinot's Palette, where the group of women drank wine, socialized and created masterpieces under the instruction of a local artist.
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Here's how sessions at Pinot's Palette typically work. A local artist teaches a group of people how to replicate a canvas painting in two or three hours. The painters can bring their own food. Wine and beer are available at the venue's bar. Music plays while guests paint and socialize.
Doggett, who worked in hospitality management and training for 20-plus years, first learned about the paint-and-sip studio concept a few years ago.
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The Doggett family opened a second Pinot's Palette location last September at 2610 Col. Glenn Highway in Fairborn, across from Wright State University. The 4,100-square-foot facility includes a black light feature in the private party room for use with florescent paint. Kim Akemon, a longtime family friend of the Doggetts, serves as the studio manager.
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For a full schedule of themed events, and more information about Pinot's Palette, visit www.pinotspalette.com/miamisburg and www.pinotspalette.com/fairborn.


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The decision to revise a facelift is a deeply personal choice. There are many reasons why someone may feel dissatisfied with a previous facelift but one of the main reasons is that they are simply unhappy or unsatisfied with the previous surgeon's work. In fact, patients with a poorly performed facelift may feel that they're in worse condition than they were before they had cosmetic surgery. The good news, however, is that many facelifts that fell short of their intended goals can be improved upon via corrective plastic surgery.
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Tom Dawes
Tom Dawes is a self-confessed 'Recovering Manufacturer'; after studying for a PhD in Supply Chain Collaboration Technologies, Tom faced a crossroads between entering the Technology or Manufacturing sectors. He chose Manufacturing, but after several successful years as an MD within the Aerospace treatments industry, the global downturn in this sector caused him to rethink. Now armed with in-depth industry insight, Tom formed Valuechain, a provider of Smart Manufacturing Software. He has developed a clear vision of how companies will collaborate in the future as integrated Valuechain's and the importance of technology to improve productivity, unlock collaboration and build intelligence.


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Hi! This is Dan Bilefsky, writing to you from the passenger seat of a tan Volvo as I travel from Quebec City to Saguenay, an industrial center in the province, on the last leg of my road trip across Quebec.
My head is spinning from the last few days in Quebec City, the stunning European-tinged capital of this province, where, among other things, I met with high school students and interviewed survivors of last year's attack on a Quebec mosque.
I also had a wine-fueled meal at L'Affaire est Ketchup, where young chefs conjure up exquisitely wrought dishes of octopus and rack of lamb on an old-style electric stove as diners gyrate to loud French pop music in between shots of Vermouth.
On my quest to understand Québécois identity, I also traveled to the sleepy farming town of Hérouxville, in the Mauricie region,, population 1,300. It was among my most memorable experiences so far for what it showed me about today's Quebec.
The nondescript town, which seems frozen in a 1950s time warp and has one set of traffic lights, a post office, a church and a dépanneur, or convenience store, ignited a provincewide culture war about a decade ago when a city counselor, André Drouin, introduced a code of conduct warning immigrants not to stone women or light them on fire. It also explained the importance of Christmas trees.
The only thing was: The town didn't have any immigrants.
The code of conduct spurred a backlash with liberals ridiculing the town as racist, closed-minded or worse. But the town's many defenders, including social conservatives, said it was merely trying to protect Quebec's way of life.
The debate was so loud that the province convened a commission on the extent to which Quebec should accommodate minorities. And so it was that I found myself shivering and walking down Hérouxville's Main Street this week in the frigid March cold, curious to see whether attitudes had shifted.
At first, the reception was equally frosty as local residents glared at me suspiciously. People who eventually talked to me did not want to give their full names.
The message, though, was clear: Even after 10 years, the town was still reeling. One man said the code was the first subject strangers brought up when he said he was from Hérouxville. A woman said the town was tired of the attention.
But another woman defended the code, saying immigrants to Quebec had to conform to local custom.
Across Quebec, on radio shows and on social media, commentators tut-tutted me for going to Hérouxville, suggesting that there were more interesting places. Others accused me of being out of touch with Quebec.
But my road trip suggests otherwise.
More than a decade later, Quebec seems consumed by some of the same questions about identity.
Late last year Quebec introduced Bill 62, a law requiring people to show their faces in order to get public services. The law has upset some Muslims, including women who wear face-covering veils known as niqabs, and say that it stigmatizes them.
This week I also met with members of the Quebec City mosque where six people were killed by a Québécois gunman more than a year ago, leading to soul-searching about the state of Islamophobia in the province. During a chat at the mosque, which is still pocked with bullet holes, its director, Mohamed Labidi, told me that he feared that the far right combined with right-wing radio was fomenting hatred against Muslims and fear of the other.
"The situation for us has gotten worse over the past decade," he said. "It has taken blood to flow for us to build bridges again."
As I travel across Quebec, I am constantly reminded how a province with a French-speaking population surrounded by an Anglophone majority in the rest of the country is understandably sensitive when it comes to preserving Quebec's culture.
Moreover, because Québécois revolted against the strictures of the Roman Catholic Church in the 1960s, secularism is deeply ingrained and outward signs of religion can cause discomfort.
The Volvo is passing snow-covered mountains as we head to Saguenay, a center for the forestry and aluminum industries. There, I want to explore how globalization and factory closings have affected a region that is among the most separatist in the province.
On a different note, I also want to sample the region's vaunted meat pies, known as tourtières!
Please follow me on my journey on Facebook, Instagram and Twitter, as well as on The Times's Reader Center. I will return with the regular Canada Letter this weekend to let you know how the rest of the trip went.


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Season of Worship:
Will You Come?
"You can't afford to pour oil out to the people at this time. The oil I have is given is for ME. I desire your worship, your time, your heart. It is TIME to refuse to give the oil I have given you. It is Time to pour out in WORSHIP, not WORD. It is TIME, in this dark hour to go forth to meet ME. I AM waiting. Will you come?
Many will say, 'Why is this waste made?' You have so much to give the people. But I say unto you, CHOOSE this day whom you will serve. Will you serve the people's needs or MINE? Will you walk by the dictates of men or by the dictates of God?
Again I say unto you, I AM waiting. Will you COME?"
Now when Jesus was in Bethany, in the house of Simon the leper, There came unto him a woman having an alabaster box of very precious ointment (or FRAGRANT OIL), and poured it on his head, as he sat at meat. But when his disciples saw it, they had indignation, saying, To what purpose is this waste?For this ointment (or FRAGRANT OIL) might have been sold for much, and given to the poor–Matthew 26:7-9, NKJV
But the hour cometh, and now is, when the true worshippers shall worship the Father in spirit and in truth: for the Father seeketh such to worship him–John 4:23
Arise My Love!
A Prophetic Word
for His Bride
"This day I call you 'Hephzibah,' for My Delight is surely in you. I have seen your seeking heart; your heart that longs after Me. I have been pleased at your insatiable desire to be with Me in a new dimension of fellowship. I will now enable you to come up higher even into the realms of My Glory, for I give unto you who seek Me eagles' wings . . . wings that will catch the wind of My Spirit; wings that will lift you into a new realm of glory.
Come unto Me, My love.
Come away from the World. Come away from the fear of man that has had you ensnared, for I have a NEW PLACE for you, a NEW ASSIGNMENT for you, a WONDERFUL BLESSING for you as you reach for the higher way. I have watched as you have been pulled back from Me by the will of man, but I say to you, the will of man will NO LONGER PREVAIL in your lives. MY WILL for your lives will NOW be accomplished. Storms of life that have dashed your hopes of a higher way are GONE this day.
You will walk with Me in glory, you will ascend to heights above that which you have dreamed, for I have given to you this day, 'Wings of Glory.'
Come to Me, My Bride!"
August 26, 1995
I want you to "ignite" hearts. Speak of Me as you know Me and others will begin to hunger and thirst after more of Me. One taste is all they will need. One spark and I will fan it. One crumb and I'll give them the whole loaf.
You are not aware of how well you know Me, but I will bring the knowledge of Me out of the depths of your heart and soul. I will "draw out" now that which I have poured into you, that which I have changed within you, that which I have increased within you that many may taste and see that I Am Good, I Am Merciful, I Am Everlasting, I Am Love, I Am Savior, I Am Blessed Forever, and the One who is the blessing for My people.
This is My work and it will simply flow from your innermost being. It will not be difficult. It will bring you great pleasure and delight.
The hearts will become inflamed with first love for Me!
This is My heart's desire.
The Bridegroom's Desire
Prophetic Word
My beloved one, My desire is for you, MY Bride, and you alone. Although My concern and compassion, My mercy, and longsufferng is for all the world and all My church, My DESIRE, My longing, and My joy is for you — My Bride.
I have seen your struggles as you have walked My way. I have seen you cast out for My name's sake. I have watched as you have spoken of Me with bridal love. I have seen the negative reaction that you have incurred. In all of this, I have watched and have seen that each negative circumstance has drawn you closer to Me. I am pleased with you. I AM pleased.
NOW, I will draw you away from negative circumstances. Now, I will seat you in heavenly places in a new level of authority with Me. Now, I will rebuke those that have criticized you. Now, I will send you as My representative to many nations and peoples to speak of Me.
You have shown Me who you are, beloved one, in many trials. Now, I will show you who I AM in a new dimension. You have known Me as Savior, as Provider, as Shepherd, and King. Now, I will reveal Myself to you, My faithful Bride, as BRIDEGROOM. I will appear to you in glory and no man will ever again take your joy from you. No one will be able to suppress you. No enemy will be able to depress you.
Come, My Bride, My desire, My joy, and crown. Come into the place I have prepared for you and you alone. Come and sit by My side. Come into this new place of fellowship. Come into a new anointing and authority. All of this I have prepared in this new day for you, My love, My dove, My undefiled.
To His Bride
Prophetic Word
Given to Penny Sheahan
On March 15, 1991
"I am standing before you now saith the Lord, desiring to cover you with My garments, to clothe you anew with My presence that My fragrance will pour forth from you. You shall no longer carry the fragrance of the world which is the stench of death, but will emit the very fragrance of life.
I see you as Ruth, lying at the feet of Boaz, longing for Me to cover you in a more intimate way than you have ever known. You long for the union of the two of us as bride and bridegroom. Nothing short of conjugal union will satisfy your heart or Mine, so I stand ready, anxiously waiting to cover you completely with My very being.
I stand at the door of your heart knocking, asking, 'Will you make yourself ready? Will you be cleansed anew with My blood? Will you put off the old garments of hate and bitterness and self-pity once and for all? Will you allow Me to purify you even as the sons of Levi that you may walk in a new and higher way in Me? (Malachi 3:3). Will you put off once and for all the sins of the flesh and be clothed in My righteousness in every area of your being that we may be One? I ask only for your willingness, and when I have your permission and know fully what you desire, I will quickly remove the old nature even as I removed the filthy garments from Joshua the High Priest. It will be a quick work of repentance I will bring to you that you may instantly be clothed with My garments (Zechariah 3:3,4 — "I will clothe you with rich robes").'
The joy you will have in this union will be unspeakable and you will fulfill My joy as you stand as MY Queen at My side clothed in the pure gold of My nature. How splendid we will be as we walk together through this earth, for surely My glory shall be yours. You shall arise and shine with My very presence — without spot or wrinkle; transparent you'll be and you shall live with Me for all eternity."
When the Dust Settles
Word of God
Thus saith the Lord: "The ungodly will flee at My presence. They shall be scattered . . . even scattered as dust before Me. For the flood of ungodly men has made My people afraid. I will NOW ARISE and DRY UP the flood of ungodliness.
I will send a mighty wind and BLOW AWAY the enemies of My people. You will look for them and not find them, for I, the Lord your God, am about to move in POWER on behalf of My people. Do not be disturbed by the things you see for you have not seen Me move in this way before. You have not seen the hills shake at My presence. You have not seen the fire that goes before Me. You have not heard the mighty wind in which I move nor seen the Way in the seas that I make as I move through the oceans of the world.
Watch as I lift you, My righteous people, above the stench of the world into the place of holiness where I dwell; above principalities and powers and their rule over you. For have I not said you will be the head and not the tail? Have I not said to them that overcome that they will rule and reign with Me? THIS is THAT TIME, saith the Lord. This is the time of the exaltation of the righteous remnant.
Again, I say unto you. WAIT. Wait through the earthquakes, wait through the fire, wait through the wind, for at the end, I will speak and you will see CLEARLY what I am asking you to do. You will see CLEARLY those of the same spirit and calling; those who will be with you in these last days. You will see CLEARLY as I lift MY remnant to rule with Me. You will see CLEARLY the character of My Son revealed in My people. Communication will be CLEAR once again. I will speak to you plainly and you will move accordingly.
Then you will see My GLORY fill the earth for you will see MY people work WITH ME in a cooperative effort to establish My Kingdom on this earth. You will see My people move in love towards one another, each one preferring the other. You shall arise and shine as one in this move and sweep VERY QUICKLY over the earth gathering all who shall be saved . . . then shall the end come, saith your God."


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Has your dentist recently mentioned that you need a root canal in Black Diamond, Washington? Root canals can seem extremely intimidating, but they really aren't as annoying as many people think. In most cases, it's the fear of the unknown that causes so much anxiety. To help put you at ease before your next dental appointment, here's what to expect from this process. Keep this information in mind for before and after your root canal in Black Diamond.
Before Your Root Canal
As you get ready for your upcoming root canal, make sure that you have your insurance information on hand. You should also bring any X-rays provided to you by your dental professional, as well as a referral card if you have one. When able, it's also a good idea to fill out any patient registration documentation ahead of time. You don't, however, need to change your eating habits prior to your appointment, unless I.V. sedation is necessary.
After Your Root Canal
After a root canal, it isn't uncommon to experience some tenderness. Sensitivity is normal during the days immediately following your procedure, especially if your tooth's roots were long. Your family dentist will probably prescribe medication, or recommend over-the-counter alternatives, to manage the pain as you heal.
Ready to Schedule Your Root Canal Procedure in Black Diamond?
Are you ready to schedule your root canal procedure in Black Diamond, Washington? Now that you know what to expect before and after your appointment, it's time to speak with a dentist. Contact Soos Creek Dental in Covington today. We'll consistently provide expert dental advice, quality service, and unbeatable results. Our team is available now to answer any and all of your questions. You can also count on our practice for cosmetic dentistry. Additionally, we commonly provide patients with dental implants and Invisalign.
Root Canal Experts in Black Diamond
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General Pediatrics and Hospital Medicine
The division of general pediatrics provides inpatient and outpatient care to all children. Our hospitalists provide care on the inpatient units of MUSC's Children's Hospital. We also care for most of the infants in the Level I Newborn Nursery.
Our faculty members' areas of expertise represent the following subspecialties, including the following:
- Asthma care
- Environmental health
- Immunizations
- International health
- Medical education Medication prescribing errors
- Medical infomatics
- Medication prescribing errors
- Obesity
- Quality of care
- School-based health care
- Toxicology
- Telehealth
Training Programs
- Academic Generalist Fellowship in Health Services Research
- Fellowship in Pediatric Hospital Medicine
- William T. Basco, M.D., MS - Division Chief
- Michelle I. Amaya M.D., M.P.H.
- David Bundy, M.D.
- Kirstin Campbell, M.D.
- Kathryn Cristaldi, M.D.
- Kristina Gustafson, M.D., MSCR
- Annie Andrews, M.D.
- Patricia G. McBurney, M.D., MSCR
- James McElligott, M.D., MS
- Claire MacGeorge, M.D., MSCR
- Sarah Mennito, M.D.
- David Mills, M.D.
- J. Routt Reigart, M.D.
- Sara Ritchie, M.D.
- James R. Roberts, M.D., MPH
- Christine San Giovanni, M.D.
- Andrea P. Summer, M.D.
- Ronald J. Teufel, II, M.D., MSCR
- Elizabeth M. Wallis M.D.
- Daniel Clay Williams, M.D.
- Conrad Williams, M.D.
Faculty with Joint Appointments in General Pediatrics
Hospitalist Services
The MUSC Pediatric Hospitalist Service is a dedicated team of highly trained academic pediatricians committed to providing the highest quality of inpatient care to your pediatric patients. At times, your child's primary care physician may request that a specially-trained pediatrician, called a hospitalist, oversee your child's care during his or her stay at the MUSC Children's Hospital.
Pediatric Hospitalists...
- evaluate patients for potential admission through the pediatric emergency department.
- admit patients, develop plans for diagnosis and treatment within 24 hours of admission.
- coordinate sub-specialty and surgical consults, as needed, while continuing to oversee the overall care of the patient.
- maintain open lines of communication with patients and their families throughout their stay and upon their discharge.
- maintain open lines of communication with patients' physicians.
- provide a detailed discharge summary for patients and their physicians.
Contact Us
MUSC Department of Pediatrics
Division of General Pediatrics
135 Rutledge Avenue, MSC 561
Charleston, SC 29425
Phone: 843-876-8512
Fax: 843-876-8709
Email: firstname.lastname@example.com


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This story still haunts Joey Logano to this day. One Halloween past, the unsuspecting Logano came home to horrifying news from his wife Brittany. They would be hitting the town for Halloween dressed as a hot dog and bun.
But the horror doesn't stop there. Because they got a bit of a jump on things, they were out to dinner in Charlotte around the same time as everyone was getting off from work. There he was, in the middle of a sea of business people in various shades of grey suits and ties… dressed as a hot dog.
Learn from poor Joey here, because he took it all in stride. He said it was a little embarrassing but it was ok, because that's just who he and his wife are: A hot dog and his bun.
Also, don't call it downtown, it's "UPtown." People from Charlotte can get weird about that.
Related: This kid transforms to be the hands down winner of best Halloween costume


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Request a Demo
Exceptional Child
c/o Scenario Learning, LLC.
2135 Dana Avenue, Suite 300
Cincinnati, OH 45207
Phone: 1.800.434.0154
Email: email@example.com
Try Our Program for Free
Thank you for your interest in the Exceptional Child Online Professional Development System. Please provide the following information and we'll be in touch soon with access details so you can preview our product at your convenience.


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The RSPCA is appealing for information after a gerbil in his cage was abandoned "like a piece of rubbish" in a skip.
A member of the public contacted the RSPCA on Tuesday (18 April) after finding the gerbil in the skip in Prescelly Close, Nuneaton.
The gerbil is now in the care of the RSPCA.
RSPCA inspector Louise Marston said: "This is so sad and it sickens me that someone just threw the gerbil in the skip to die, as though he was a piece of rubbish.
"He had water in his bottle, but the cage in general was quite dirty. It is pretty clear that whoever did this did so knowing that the gerbil was in there.
"If he hadn't have been found, he would have died a horrible death. The skip wasn't even full, so a lot of extra rubbish would have been thrown on top of this poor gerbil, if he hadn't have been found. Thankfully he is recovering now – he is very lucky."
Anyone with information about who abandoned the gerbil should contact the RSPCA on 0300 123 8018, and leave a message for Inspector Marston.
"If someone gets a pet then they need to realise that they have to commit to being responsible for the animal throughout their life," added Inspector Marston. "It is terrible to think that someone thought they could just through the gerbil away because they no longer wanted him."
Last month, the RSPCA released its annual Cruelty Statistics, which showed that animal cruelty complaints across England and Wales had increased by 3.5%. In Warwickshire, the number of animal cruelty complaints inspectors investigated in 2016 was 1,307 – up from 1,280 in 2015 and an increase of 2.1%.
Dermot Murphy, Assistant Director of the RSPCA Inspectorate, said: "It never fails to shock me when I look back on the extreme instances of animal cruelty the RSPCA has been called upon to investigate.
"It continues to outrage and sadden me that people can be capable of such deliberate brutality towards animals, but equally it drives me on to ensure that perpetrators of animal cruelty are put before the courts.
"I believe that the figures from last year show that we're not becoming crueler, but that people are simply less willing to stand by and do nothing if they think an animal is suffering.
"People are increasingly likely to share images or footage on their social media accounts of animals they believe are not being cared for properly, while many will see material their friends have shared and then contact us about them."
The RSPCA is a charity and we rely on public donations to exist. To assist our inspectors in carrying out their vital work please text HELP to 78866 to give £3. (Texts cost £3 + one standard network rate message.)


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Friday, April 10, 2020
Palestinian artist Fouad Agbaria opens latest exhibition: Weaving and Cracks
Eli Lillis
- September 19, 2018
Main News
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Foud Agbaria is
as 'one of the most prominent young Palestinian artists working today'. In Agbari's Umm Al-Fahem studio, work for his newest collection Weaving and Cracks began more than one year ago, when he started studying the carpets of the Arab world and the Middle East. Painting then began and in seven months nearly 30 works were completed for this series.
Fouad Agbaria's 'Weaving and Cracks' exhibit at Zahyeh Gallery.
During the opening of his latest exhibition at Zawyeh Gallery in Ramallah, Agbaria described his inspiration.
"I am not divorced from the reality around me. When I am painting my country, villages, hills, flora, all of my works are political in their intent. In reality I'm painting the wars, the walls, and the occupation," Agbaria told Palestine Monitor.
Two works from the 'Weaving and Cracks' collection.
In this work, the recurrent theme of the carpet stands as metaphor for Islam, the Arab world and the artists own youthful, nostalgia for home. Technically, work on the carpet is characterized by harmonic and complex sequences, yet this harmony contains many deliberate and spontaneous errors, jarring the viewer.
Juxtaposed into the traditional Arabic carpet motifs are various symbols, signifying childhoods ruined by wars, and tools of killing and destruction, which have become part of the topography of the Arab world.
The front of Zawyeh Gallery.
In this contrast, the artist creates a domestic space speaking of comfort and familiarity, yet then inserts strange and threatening motifs, further distressing the viewer.
Speaking about the role of the artists in Palestinian society, Agbaria said; "There is no doubt that what the Palestinian artist produces will be affected by what is happening around them, even if the painting is only of a bird or a rose."
Viewers mingle in front of two pieces from the series 'weaving and cracks.'
"A lifetime of violence and occupation are psychologically and physically reflected in every brushstroke and layering of colour. Artists have an important role in documenting the suffering of the self and the community, and through this artistic work an artist can convey a message to the world, to tell the suffering of the people."
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5 bedroomed, detached, country home available for rent all year round. Set in a picturesque valley 20km south of Chablis, Burgundy, France. The house is set up for a relaxing, self catering holiday. Quiet village setting with boulangerie close by.
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Michael Dill Photography has two locations, one in Jupiter, Florida and the other in Robbinsville, NJ.
Our locations offer Lifestyle Photography. Each session is custom designed to produce an heirloom piece that your family will want to hang in that prominent location in your home. All wall portraits can be custom framed and/painted.
Thinking about a beach or family session. Michael will guide you in the proper clothing to wear and appropriate time to produce that special image.
Business headshots are available on location. Our portable studio setup can come to your location. We provide both full size and web size digital files that you can use for advertising and on the web.
Michael has been photographing action sports for over 25 years from the youth leagues to the major leagues. Need action photos of your child? Call and schedule an appointment where Michael will focus on your individual child. Does your team want action photos, banners, custom collages or team and individual photos? Michael Dill Photography is your choice for unsurpassed quality.
Our weddings offer two photographers. We photograph traditional and photojournalist style weddings. Wedding packages include a disk of images or the traditional packages with wedding albums. Engagement sessions are also available.
Look at our special ballerina pieces. We custom design each session and Michael is well-known for his ballerina designs. Call today to schedule your session.
Our landscape photos are available in sizes ranging from 8x10 to 30x40. Please email us for specific pricing on landscape photos.


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As a consultant for Regeneration Works: Places of Faith, Stephen works with two national organizations; Faith and the Common Good, and The National Trust of Canada, and a team of experts to help faith buildings from coast to coast become more vibrant through a better understanding of their buildings. By understanding the building better, it becomes easier to maximize it's usage and impact on both the faith community and the community at large.
Regeneration Works offers four core services to faith-based organizations and their building stewards:
Building audits help determine:
The Regeneration Audit will help faith communities who may be under considerable financial or community pressure to restructure, amalgamate, etc. This walk-through audit acts as an outside set of eyes and viewpoint to ensure the best consensus decisions can be made moving forward for all involved. Bringing a triple-bottom-line sustainability perspective (environmental, social, financial), this audit will help participants evaluate issues such as building, operations, energy, historical concerns, community activity within and beyond the walls, and how to engage the community to strengthen the sustainability goals moving forward. Natural Resources Canada (NRCan)'s Energy Star Portfolio Manager is used in this walk-through audit as a means to compare buildings in question. Discounts will apply if you would like this audit to encompass more than one faith community.
By working with our team of professionals during your faith community's time of need, we will guide you with the best unbiased information possible, to help you make the best decision for the future of your community and beyond. The Regeneration Audit can be delivered by itself or as part of a comprehensive consultation with your faith community.
Planning, strategy and logistics in the selection of building partners and collaborating groups.
Support the development of networks and plan cooperative efforts to bring community members and local organizations into spaces of worship to regenerate and support the space.
Regeneration works occasionnaly organizes and hosts workshops on supporting and revitalizing spaces of worship and community hubs.


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Dr. David Eagleman journeys into the future and asks what's next for the human brain, and for our species. He reveals that in the future our descendants may be so different to us that we will be strangers to them.Premiere Date: November 18, 2015 | Runtime: 00:54:41
Neuroscientist David Eagleman explores the human brain in an epic series that reveals the ultimate story of us, why we feel and think the things we do. This ambitious project blends science with innovative visual effects and compelling personal stories, and addresses some big questions. By understanding the human brain, we can come close to understanding humanity.


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Hi there!!
Welcome to my Cuba Blog! My name is Flavia and I'll be using my words and tremendous passion for traveling to inspire others to discover the wonders Cuba has to offer. I was born in the countryside of Cuba and moved to Havana when I was 6 years old. I lived in there until 2009 when my parents moved my brother and I 90 miles north to the sunshine state – Florida. Adapting to a whole new language and culture was extremely challenging and still is. Leaving friends and family behind is not the easiest thing to do and takes time and effort to move past the tears and start a new life. I love America, I love my life here and the many opportunities I've had. There is nothing like having freedom run through your body and proudly call yourself an American; but I will never forget where I came from. I take Cuba with me, every single day, every minute, everywhere I go. There is nothing I love more than talking to people about my country and culture. Recommending where to go, what to eat or what to see to people who are planning on visiting is my drug. Cuba is a small country filled with kind, humble, loving people. It's a step back in time where technology has not taken over yet. It's a country where perseverance and dreams make people do the impossible. A country so full of natural beauty that every street, every corner, every building is worth photographing. I want to take you in the adventure of your life. From the crowded streets of Havana to the calm country side of Santiago; come explore this magnificent island alongside me.
I am not the same, having seen the moon shine on the other side of the world. — Gustave Flaubert