interacts_with DB00477?
Epigenetic regulation of protein tyrosine phosphatase Q05209 in triple-negative breast cancer . AIMS : The present study showed that the expression of Q05209 is epigenetically regulated . DB00928 ( 5-Azac ) , a DNA hypomethylating agent , significantly increased the expression of Q05209 at low concentrations ( 1μM and 2.5μM ) and decreased the expression of Q05209 at 5μM in the MDA-MB-231 and BT-549 triple-negative breast cancer cell lines . MAIN METHODS : Human MCF-7 , MDA-MB-231 and BT-549 cells were exposed to different concentrations of 5-Azac for 24 and 48h . RT-PCR was performed to determine the mRNA expression of Q05209 , P12830 and miRNA-124 . Western blotting was performed to assess the protein expression of various proteins , including Q05209 , P12830 , P26358 and PARP . KEY FINDINGS : 5-Azac , a DNA hypomethylating agent , significantly increased the expression of Q05209 at low concentrations ( 1μM and 2.5μM ) and decreased Q05209 expression at 5μM . We provide the first evidence that Q05209 expression is epigenetically regulated and that it is up-regulated at a lower dose of a P26358 inhibitor in MDA-MB-231 and BT-549 cells . Interestingly , the levels of miRNA-124 were increased only at 5μM , the concentration at which Q05209 expression was suppressed . SIGNIFICANCE : To the best of our knowledge , this is the first report that highlights the therapeutic potential of low-dose 5-Azac for the treatment of TNBC . Therefore , 5-Azac , an agent that has already been tested in acute myeloid leukemia , may be more effective at lower doses for the treatment of triple-negative breast cancer . Real-time DNA binding measurements of the P14921 recombinant oncoproteins reveal significant kinetic differences between the Q8NFH3 and Q9H3D4 isoforms . The sequence-specific DNA binding of recombinant Q8NFH3 and Q9H3D4 P14921 oncoprotein was examined quantitatively to determine whether the loss of the Exon VII phosphorylation domain in Q8NFH3 P14921 or the phosphorylation of expressed Exon VII in Q9H3D4 P14921 had an effect on DNA binding activity . The kinetics of sequence-specific DNA binding was measured using real-time changes in surface plasmon resonance with BIAcore ( registered trademark , Pharmacia Biosensor ) technology . The real-time binding of Q8NFH3 and Q9H3D4 P14921 displayed significant differences in kinetic behavior . Q9H3D4 P14921 is characterized by a fast initial binding and conversion to a stable complex , whereas Q8NFH3 P14921 exhibits a slow initial binding and conversion to a stable complex . All of the Q9H3D4 P14921 DNA binding states are characterized by rapid turnover , whereas the Q8NFH3 P14921 DNA binding states are 4-20 times more stable . A model describing these kinetic steps is presented . Stoichiometric titrations of either Q8NFH3 or Q9H3D4 P14921 with specific oligonucleotides show 1:1 complex formation . The DNA sequence specificity of the Q8NFH3 and Q9H3D4 P14921 as determined by mutational analysis was similar . The in vitro phosphorylation of Q9H3D4 P14921 by P62158 kinase II obliterates its binding to specific DNA , suggesting that the regulation of Q9H3D4 P14921 sequence-specific DNA binding occurs through phosphorylation by a calcium-dependent second messenger . The Q8NFH3 P14921 lacks this regulatory domain ( Exon VII ) , and binding to its specific DNA sequence is not sensitive to calcium signaling . Poly(ADP-ribose) metabolism in X-irradiated Chinese hamster cells : its relation to repair of potentially lethal damage . DB02701 -adenine dinucleotide ( NAD+ ) is the substrate used by cells in poly(ADP-ribose) synthesis . X-irradiation of log-phase Chinese hamster cells caused a rapid decrease in NAD+ levels which was linearly dependent on radiation dose . The activity of ADP-ribosyl transferase ( P09874 ) also increased linearly with radiation dose . The decrease of NAD+ was slower , and the increase in P09874 activity was less pronounced , in a radiation sensitive line , V79- AL162 /S-10 . An inhibitor of P09874 , m-aminobenzamide , largely prevented the depletion of cellular NAD+ and reduced the rate at which P09874 activity disappeared during post-irradiation incubation . Post-irradiation treatment with hypertonic buffer or with medium containing D2O -- which inhibit repair of radiation-induced potentially lethal damage -- enhanced the depletion of NAD+ and prevented the reduction in P09874 activity following irradiation . The characteristics of the effects of treatment with hypertonic buffer on NAD+ metabolism were qualitatively similar to the effects that such treatment has on radiation-induced cell killing . These results suggest that poly(ADP-ribose) synthesis after irradiation plays a role in the repair of potentially lethal damage . DB04873 ( SB 207266 ) , a selective Q13639 receptor antagonist , reduces serotonin potentiation of neurally-mediated contractile responses of human detrusor muscle . The aim of this study is to evaluate the potency of piboserod ( SB 207266 ) , a selective 5-HT(4) receptor antagonist , at inhibiting the 5-HT(4)-mediated potentiating effect of serotonin ( 5-HT ) on the neurally-mediated contractile responses of human detrusor strips to electrical field stimulations ( O43281 ) . Strips of human detrusor muscle were mounted in Krebs-HEPES buffer under a resting tension of 500 mg and O43281 ( 20 Hz , 1 ms duration at 300 mA for 5 s ) was applied continuously at 1 min intervals . After stabilization of the O43281 -induced contractions , concentration-response curves to 5-HT ( 0.1 nM-100 microM ) were constructed in the absence or presence of 1 or 100 nM of piboserod . The experiments were performed in the presence of methysergide ( 1 microM ) and ondansetron ( 3 microM ) to block 5HT(1)/5HT(2) and 5-HT(3) receptors , respectively . 5-HT potentiated the contractile responses to O43281 of human bladder strips in a concentration-dependent manner , with a maximum mean of 60.0+/-19.9 % of the basal O43281 -evoked contractions . DB04873 did not modify the basal contractions but concentration-dependently antagonized the ability of 5-HT to enhance bladder strip contractions to O43281 . In presence of 1 and 100 nM of piboserod , the maximal 5-HT-induced potentiations were reduced to 45.0+/-7.9 and 38.7+/-8.7 % , respectively . A mean apparent antagonist dissociation constant value ( K(B) ) of 0.56+/-0.09 nM was determined . These data show the ability of piboserod to antagonize with high potency the enhancing properties of 5-HT on neurally-mediated contractions of isolated human bladder strips . Therefore , the 5-HT(4) receptor might represent an attractive pharmacological target for the treatment of overactive bladder . Isolation and characterization of P62333 . A novel ATPase family component of the yeast 26 S proteasome . Using a genetic strategy designed to find proteins involved in the function of the Saccharomyces cerevisiae transcriptional activator GAL4 , we isolated mutants in two genes which rescue a class of gal4 activation domain mutants . One of these genes , P62195 , encodes a member of a large family of putative ATPases , the Conserved ATPase containing Domain ( CAD ) proteins ( also known as AAA proteins ) that are involved in a wide variety of cellular functions . Subsequently , P62195 was identified as a subunit of the 26 S proteasome . We have now cloned the gene defined by the second complementation group . P62333 encodes an essential 49-kDa protein that is also a member of the CAD family and is 43 % identical to P62195 . The mutation in sug2-1 , like that in sug1-1 , is found in the CAD near the highly conserved ATPase motif . We present biochemical and genetic evidence that P62333 is associated in vivo with P62195 and is a novel P27708 subunit of the 26 S proteasome . With its highly conserved mammalian homologs , human Q8NFH3 and ground squirrel CADp44 , P62333 defines a new class of proteasomal CAD proteins . 5-hydroxytryptamine and its receptors in systemic vascular walls . 5-hydroxytryptamine ( 5-HT ) in the bloodstream is largely contained in platelets and circulates throughout the entire vascular system . 5-HT released from activated platelets dramatically changes the function of vascular smooth muscle cells ( VSMCs ) and endothelial cells ( ECs ) . In VSMCs , 5-HT induces proliferation and migration via 5- Q13049 receptors . These effects are further enhanced by vasoactive substances such as thromboxane A2 and angiotensin II . 5- Q13049 receptor activation in VSMCs also causes both enhancement of prostaglandin I2 production by inducing cyclooxygenase-2 and reduction of nitric oxide ( NO ) by suppressing inducible NO synthase . Evidence showing that 5-HT in ECs plays a principal role in angiogenesis now exists . Stimulation of 5-HT1 and/or 5-HT2 receptors has been implicated in the angiogenic effect of 5-HT . The extracellular signal-regulated kinase and endothelial NO synthase ( P29474 ) activation-dependent pathways are involved in the mechanisms . Moreover , Q13639 receptors in ECs have been shown to also regulate angiogenesis . Recent reports show sarpogrelate , a selective antagonist of the 5- Q13049 receptor , indirectly enhances the function of P28222 receptors in ECs via inhibition of 5- Q13049 receptors in VSMCs or platelets . This indirect action of P28222 receptors in ECs may increase NO production derived from P29474 and a vasodilator response . Furthermore , sarpogrelate and other 5- Q13049 receptor antagonists have been shown to reduce the constitutive activity of 5- Q13049 receptors . It is believed that increasing evidence on the role of 5-HT receptors will contribute to the expansion of the clinical application of existing therapeutic drugs such as sarpogrelate , and to the development of new 5-HT receptor-related drugs for treating cardiovascular diseases . Cell adhesion molecule uvomorulin expression in human breast cancer cell lines : relationship to morphology and invasive capacities . Loss of cell-cell adhesion in carcinoma cells may be an important step in the acquisition of an invasive , metastatic phenotype . We have examined the expression of the epithelial-specific cell adhesion molecule uvomorulin ( P12830 , cell- P12830 , L- P62158 ) in human breast cancer cell lines . We find that fibroblastoid , highly invasive , vimentin-expressing breast cancer cell lines do not express uvomorulin . Of the more epithelial-appearing , less invasive , keratin-expressing breast cancer cell lines , some express uvomorulin , and some do not . We examined the morphologies of the cell lines in the reconstituted basement membrane matrix Matrigel and measured the ability of the cells to traverse a Matrigel-coated filter as in vitro models for detachment of carcinoma cells from neighboring cells and invasion through basement membrane into surrounding tissue . Colonies of uvomorulin-positive cells have a characteristic fused appearance in Matrigel , whereas uvomorulin-negative cells appear detached . Cells which are uvomorulin negative and vimentin positive have a stellate morphology in Matrigel . We show that uvomorulin is responsible for the fused colony morphology in Matrigel since treatment of uvomorulin-positive MCF-7 cells with an antibody to uvomorulin caused the cells to detach from one another but did not induce invasiveness in these cells , as measured by their ability to cross a Matrigel-coated polycarbonate filter in a modified Boyden chamber assay . Two uvomorulin-negative , vimentin-negative cell lines are also not highly invasive as measured by this assay . We suggest that loss of uvomorulin-mediated cell-cell adhesion may be one of many changes involved in the progression of a carcinoma cell to an invasive phenotype . Nitrergic response to cyclophosphamide treatment in blood and bone marrow . Daily intraperitoneal injection of cyclophosphamide ( P15085 ) ( 50 mgkg(-1) of body weight ) for 5 days resulted in reduced levels of marrow and blood cellularity , which was most pronounced in 18 days post-treatment ( pt ) . On day 18 after P15085 treatment the enhancedlevels of nitric oxide ( NO ) precursors and metabolites ( L-arginine , L-citrulline , reactive nitrogen species ( RNS ) ) of marrow and blood cells ( platelet , neutrophil , lymphocyte and monocyte ) resulted from up-regulation of Ca(II)/calmodulin( P62158 )-independent " inducible " NO synthase ( P35228 ) , with a lessercontribution of Ca(II)/ P62158 -dependent " constitutive " P29474 isoforms to systemic NO.Biphasic response to P15085 of marrow nitrergic system , i.e. both P35228 and P29474 showed significantly depressed activities , as well as diminished levels of NO metabolites on day 9 pt , suggested that signals in addition to NO might be involved in P15085 -induced inhibition of hematopoesis , while a gradual increase of neutrophil and platelet NOS activity appeared to be contributed to a P15085 -induced development of granulopenia , thrombocytopenia and hemorrhage . Exploring schizophrenia drug-gene interactions through molecular network and pathway modeling . In this study , we retrieved 39 schizophrenia-related antipsychotic drugs from the DrugBank database . These drugs had interactions with 142 targets , whose corresponding genes were defined as drug targeted genes . To explore the complexity between these drugs and their related genes in schizophrenia , we constructed a drug-target gene network . These genes were overrepresented in several pathways including : neuroactive ligand-receptor pathways , glutamate metabolism , and glycine metabolism . Through integrating the pathway information into a drug-gene network , we revealed a few bridge genes connected the sub-networks of the drug-gene network : Q12879 , O60391 , Q14957 , Q13224 , P21728 , and P14416 . These genes encode ionotropic glutamate receptors belonging to the DB01221 receptor family and dopamine receptors . DB00502 was the only drug to directly interact with these pathways and receptors and consequently may have a unique action at the drug-gene interaction level during the treatment of schizophrenia . This study represents the first systematic investigation of drug-gene interactions in psychosis . [ Regulation of P04271 expression during long term potentiation ] . In this study , contributions of intracellular regulatory cascades in the induction of P04271 expression in rat hippocampal P00915 area during long term posttetanic potentiation ( LTP ) were estimated . The activation of transcription factor p53 ( positive regulator of P04271 transcription ) by nutlin-3 increased the basal content of P04271 mRNA up to 151 % of the control level , which was significantly lower than its content in tetanized slices ( 280 % ) . Therefore , p53 seems to be not unique transcription factor upregulating P04271 expression during LTP . The inhibitor of Ca2+/calmodulin-dependent kinases ( CaMKs ) KN-93 fully blocked the increase of P04271 mRNA after tetanization , while KN-92 ( inactive analogue of KN-93 ) was ineffective . The inhibitor of CaMKII and receptor tyrosine kinases DB02152 essentially suppressed P04271 expression during LTP , the inhibition of MAPK p38 or P51812 moderately decreased , and the inhibition of Q02750 did not influence P04271 mRNA content . Thus , CaMKs play a key role in the induction of P04271 expression during LTP . [ Functional characteristics of calcium-sensitive adenylyl cyclase of ciliate Tetrahymena pyriformis ] . DB01373 -sensitive forms of adenylyl cyclase ( AC ) were revealed in most vertebrates and invertebrates and also in some unicellular organisms , in particular ciliates . We have shown for the first time that calcium cations influence the AC activity of ciliate Tetrahymena pyriformis . These cations at the concentrations of 0.2-20 microM stimulated the enzyme activity , and maximum of catalytic effect was observed at 2 microM Ca2+ . DB01373 cations at a concentrations of 100 microM or higher inhibited the AC activity . P62158 antagonists W-5 and W-7 at the concentrations of 20-100 microM inhibited the catalytic effect induced by 5 microM Ca2+ and blocked the effect at higher concentrations of Ca2+ . DB00477 , another calmodulin antagonist , reduced Ca2+-stimulated AC activity only at the concentrations of 200-1000 microM . AC stimulating effects of serotonin , P01133 and DB02527 increased in the presence of 5 microM Ca2+ . AC stimulating effects of P01133 , DB02527 and insulin decreased in the presence of 100 microM Ca2+ , and AC stimulating effect of DB02527 decreased also in the presence of calmodulin antagonists ( 1 mM ) . At the same time , stimulating effect of D-glucose in the presence of Ca2+ and calmodulin antagonists did not change essentially . The data obtained speak in favor of the presence of calcium-sensitive forms of AC in ciliate T. pyriformis which mediate enzyme stimulation by P01133 , DB02527 , insulin , and serotonin . Autonomous CaMKII requires further stimulation by Ca2+/calmodulin for enhancing synaptic strength . A hallmark feature of Ca(2+)/calmodulin ( P62158 ) -dependent protein kinase II ( CaMKII ) is generation of autonomous ( Ca(2+)-independent ) activity by T286 autophosphorylation . Biochemical studies have shown that " autonomous " CaMKII is ∼5-fold further stimulated by Ca(2+)/ P62158 , but demonstration of a physiological function for such regulation within cells has remained elusive . In this study , CaMKII-induced enhancement of synaptic strength in rat hippocampal neurons required both autonomous activity and further stimulation . Synaptic strength was decreased by CaMKIIα knockdown and rescued by reexpression , but not by mutants impaired for autonomy ( T286A ) or binding to DB01221 -type glutamate receptor subunit 2B ( Q13224 ; formerly Q13224 ; I205K ) . Q8N1N2 rescue was seen with constitutively autonomous mutants ( T286D ) , but only if they could be further stimulated ( additional T305/306A mutation ) , and not with two other mutations that additionally impair Ca(2+)/ P62158 binding . Compared to rescue with wild-type CaMKII , the P62158 -binding-impaired mutants even had reduced synaptic strength . One of these mutants ( T305/306D ) mimicked an inhibitory autophosphorylation of CaMKII , whereas the other one ( Δstim ) abolished P62158 binding without introducing charged residues . Inhibitory T305/306 autophosphorylation also reduced Q13224 binding , but this effect was independent of reduced Ca(2+)/ P62158 binding and was not mimicked by T305/306D mutation . Thus , even autonomous CaMKII activity must be further stimulated by Ca(2+)/ P62158 for enhancement of synaptic strength . DB00169 down-regulates monocyte TLR expression and triggers hyporesponsiveness to pathogen-associated molecular patterns . Toll-like receptors ( TLR ) represent an ancient front-line defence system that enables the host organism to sense the presence of microbial components within minutes . As inducers of inflammation , TLR act as important triggers of distinct entities such as sepsis or autoimmune disease exacerbation . We report here that vitamin D3 [ 1alpha,25-dihydroxycholecalciferol , 1,25(OH)(2)D3 ] suppresses the expression of O60603 and O00206 protein and mRNA in human monocytes in a time- and dose-dependent fashion . Despite 1,25(OH)(2)D3-induced up-regulation of P08571 , challenge of human monocytes with either LPS or lipoteichoic acid resulted in impaired P01375 and procoagulatory tissue factor ( CD142 ) production , emphasizing the critical role of TLR in the induction of inflammation . Moreover , reduced TLR levels in 1,25(OH)(2)D3-treated phagocytes were accompanied by impaired NF-kappaB/RelA translocation to the nucleus and by reduced p38 and Q8NFH3 /44 ( extracellular signal-regulated kinase 1/2 ) phosphorylation upon TLR-ligand engagement . Both TLR down-regulation and P08571 up-regulation were substantially inhibited by the vitamin D receptor ( P11473 ) antagonist ZK 159222 , indicating that the immunomodulatory effect of 1,25(OH)(2)D3 on innate immunity receptors requires P11473 transcription factor activation . Our data provide strong evidence that 1,25(OH)(2)D3 primes monocytes to respond less effectively to bacterial cell wall components in a P11473 -dependent mechanism , most likely due to decreased levels of O60603 and O00206 . DB00227 -stimulated superinduction of P16581 , P05362 and P19320 in P01375 activated human vascular endothelial cells . Inhibitors of P04035 ( statins ) reveal important pharmacological effects in addition to reducing the plasma LDL cholesterol level . In the pathogenesis of arteriosclerosis , transendothelial migration of various leukocytes including monocytes is a crucial step . We , therefore , investigated the expression of P16581 , intercellular cell adhesion molecule-1 ( P05362 ) and vascular cell adhesion molecule-1 ( P19320 ) in vascular endothelial cells as influenced by lovastatin . Human umbilical vein endothelial cells ( HUVECs ) express significant amounts of selectins and cell adhesion molecules ( CAMs ) within a few hours after stimulation with P01375 . This effect is potentiated by 100-200 % when the cells are pretreated with 0.1-2.5 microM lovastatin . The lovastatin-mediated increase in the cytoplasm and at the cell surface is dose-dependent and significant at lovastatin concentrations comparable to plasma levels in patients under lovastatin treatment . The lovastatin-potentiated increase of P16581 and CAMs is correlated with a corresponding increase of selectin- and P62158 -specific mRNA . We conclude that , in vivo , statin treatment could trigger an enhanced recruitment of macrophages that might support the cholesteryl ester efflux from the arteriosclerotic plaque . The cloning and reintroduction into animal cells of a functional CAD gene , a dominant amplifiable genetic marker . Rodent cells resistant to DB03459 , a specific inhibitor of the aspartate transcarbamylase activity of the multifunctional P27708 , overproduce CAD as a result of amplification of the CAD gene . We cloned a functional CAD gene from Syrian hamster cells using a cosmid vector . Two independently isolated cosmids containing CAD genes have inserts 40 and 45 kb long . We introduced the cloned genes into CAD-deficient Chinese hamster ovary ( CHO ) cell mutants by fusing them with protoplasts of Escherichia coli containing the cosmids . We also introduced the cloned genes into wild-type CHO cells by selecting cells that became resistant to high concentrations of DB03459 following protoplast fusion . The transformants of the mutant and wild-type CHO cells contain multiple active copies of the donated Syrian hamster CAD genes . The cloned genes in three independent transformants are integrated into host-cell chromosomes at single locations identified by in situ hybridization . In two of these transformants , the genes are located in one X chromosome or in a chromosome resembling the X . In the third case , the genes are located in a small metacentric or rearranged chromosome . Mechanism of inhibition of the P42262 AMPA receptor channel opening by talampanel and its enantiomer : the stereochemistry of the 4-methyl group on the diazepine ring of 2,3-benzodiazepine derivatives . Stereoselectivity of 2,3-benzodiazepine compounds provides a unique way for the design of stereoisomers as more selective and more potent inhibitors as drug candidates for treatment of the neurological diseases involving excessive activity of AMPA receptors . Here we investigate a pair of enantiomers known as DB04982 and its ( + ) counterpart about their mechanism of inhibition and selectivity toward four AMPA receptor subunits or P42261 -4 . We show that DB04982 is the eutomer with the endismic ratio being 14 for the closed-channel and 10 for the open-channel state of P42262 . Kinetic evidence supports that DB04982 is a noncompetitive inhibitor and it binds to the same site for those 2,3-benzodiazepine compounds with the C-4 methyl group on the diazepine ring . This site , which we term as the " M " site , recognizes preferentially those 2,3-benzodiazepine compounds with the C-4 methyl group being in the R configuration , as in the chemical structure of DB04982 . Given that DB04982 inhibits P42261 and P42262 , but is virtually ineffective on the P42263 and P48058 AMPA receptor subunits , we hypothesize that the " M " site(s) on P42261 and P42262 to which DB04982 binds is different from that on P42263 and P48058 . If the molecular properties of the AMPA receptors and DB04982 are used for selecting an inhibitor as a single drug candidate for controlling the activity of all AMPA receptors in vivo , DB04982 is not ideal . Our results further suggest that addition of longer acyl groups to the N-3 position should produce more potent 2,3-benzodiazepine inhibitors for the " M " site . Salmon DB00644 and its analogues bind the human placental receptor . OBJECTIVE : The presence of DB00644 receptors in the human placenta has been recognized for a number of years . However , mammalian DB00644 , which is expressed in placental tissues , has limited affinity for the chorionic receptor . On the basis of immunological and bioactivity data , we have previously proposed that the chorionic DB00644 may differ from mammalian DB00644 . METHODS : We have studied the affinity of another isoform of DB00644 ( ie , salmon DB00644 and stable analogues of this DB00644 isoform ) , and compared their receptor affinity to that of mammalian DB00644 and its analogues . RESULTS : Using our receptor assay method with the labeled mammalian DB00644 analogue DB06719 , salmon DB00644 had a twofold greater affinity for the placental P30968 than did mammalian DB00644 and for the stable salmon DB00644 analogue the affinity was increased tenfold . Using a homologous receptor assay method with a stable salmon DB00644 analogue as label , the affinity for this salmon DB00644 analogue had a K(d) of 101 nmol/L . CONCLUSION : The presence of these higher affinity receptors for non-mammalian DB00644 in the human placenta has led us to propose that the chorionic tissues may express more than one isoform of DB00644 and that non-mammalian DB00644 , such as salmon DB00644 , may be potent regulators of placental functions . Effect of growth factors on nuclear and mitochondrial ADP-ribosylation processes during astroglial cell development and aging in culture . Epidermal growth factor ( P01133 ) , basic fibroblast growth factor ( P09038 ) , insulin-like growth factor-I ( P05019 ) and insulin ( P01308 ) are powerful mitogens and may regulate gene expression in cultured astrocytes by ADP-ribosylation process . Nuclear poly-ADP ribose polymerase ( PARP ) and mitochondrial monoADP-ribosyltransferase ( P09874 ) are the key enzymes involved in poly-ADP-ribosylation and mono ADP-ribosylation , respectively . In this investigation the effect of P01133 , P09038 , P05019 or P01308 on nuclear PARP and mitochondrial P09874 activities were assessed in nuclei and mitochondria purified from developing ( 30 DIV ) or aging ( 90 and 190 DIV ) primary rat astrocyte cultures . A marked increase of PARP activity in P09038 or P05019 treated astroglial cell cultures at 30 DIV was found . Nuclear PARP and mitochondrial P09874 activities were greatly stimulated by treatment with P01133 or P01308 alone or together in astrocyte cultures at 30 DIV . Nuclear PARP and mitochondrial P09874 activities showed a more remarkable increase in control untreated astrocyte cultures at 190 DIV than at 90 DIV . These findings suggest that ADP-ribosylation process is involved in DNA damage and repair during cell differentiation and aging in culture . Twelve hours treatment with P01133 , P01308 or P09038 significantly stimulated nuclear PARP and mitochondrial P09874 activities in 190 DIV aging astrocyte cultures . The above results indicate that P01133 , P01308 and P09038 may play a crucial role in the post-translational modification of chromosomal proteins including ADP-ribosylation process in in vitro models . This suggests that growth factors regulate genomic stability in glial cells during development and maturation , stimulating nuclear and mitochondrial ADP-ribosylation processes in developing or aging astrocyte cultures . P04271 -mediated inhibition of the phosphorylation of P14136 is prevented by TRTK-12 . P04271 belongs to a family of calcium-binding proteins involved in cell cycle and cytoskeleton regulation . We observed an inhibitory effect of P04271 on glial fibrillary acidic protein ( P14136 ) phosphorylation , when stimulated by DB02527 or Ca2+/calmodulin , in a cytoskeletal fraction from primary astrocyte cultures . We found that P04271 has no direct effect on P62158 KII activity , the major kinase in this cytoskeletal fraction able to phosphorylate P14136 . The inhibition of P14136 phosphorylation is most likely due to the binding of P04271 to the phosphorylation sites on this protein and blocking the access of these sites to the protein kinases . This inhibition was dependent on Ca2+ . However , DB01593 could substitute for Ca2+ . The inhibitory effect of P04271 was prevented by TRTK-12 , a peptide that blocks P04271 interaction with several target proteins including glial fibrillary acidic protein . These data suggest a role for P04271 in the assembly of intermediate filaments in astrocytes . Cav1.2 L-type Ca²⁺ channels mediate cocaine-induced P42261 trafficking in the nucleus accumbens , a long-term adaptation dependent on ventral tegmental area Ca(v)1.3 channels . AMPA receptor ( AMPAR ) plasticity at glutamatergic synapses in the mesoaccumbal dopaminergic pathway has been implicated in persistent cocaine-induced behavioral responses ; however , the precise mechanism underlying these changes remains unknown . Utilizing cocaine psychomotor sensitization , we have examined phosphorylation of P42261 at key residues serine 845 ( S845 ) and S831 , as well as P42261 cell surface levels in the nucleus accumbens ( NAc ) of cocaine-preexposed mice and the role of brain-specific Ca(v)1.2 and Ca(v)1.3 L-type Ca²⁺ channels ( LTCCs ) , therein . We found higher basal levels of S845 phospho- P42261 ( P- P42261 ) and cell surface P42261 in the NAc following protracted withdrawal from cocaine exposure , changes that occur independently of LTCCs . In contrast , we found that a cocaine challenge that elicits expression of the cocaine-sensitized response increases S831 P- P42261 that further increases surface P42261 beyond the higher basal levels . Intra-NAc pharmacological manipulations indicate that the Ca(v)1.2-activated P62158 kinase II ( CaMKII ) mediates cocaine-induced increase in S831 P- P42261 and that both Ca(v)1.2-activated CaMKII and extracellular signal-regulated kinase 2 ( P28482 ) mediate the increase in P42261 cell surface levels specific to the sensitized response . Experiments using adenoassociated viral vectors expressing Ca(v)1.3 and P28482 siRNA further indicate that recruitment of the Ca(v)1.2 pathway in the NAc is dependent on ventral tegmental area Ca(v)1.3 LTCCs and P28482 . Together , these results identify candidate pathways that mediate cocaine-induced AMPAR plasticity in the NAc and provide a mechanism linking LTCCs and P42261 plasticity to cocaine-induced persistent behavioral changes . Dependence of gonadotropin-releasing hormone-induced neuronal MAPK signaling on epidermal growth factor receptor transactivation . The hypothalamic decapeptide , gonadotropin-releasing hormone ( DB00644 ) , utilizes multiple signaling pathways to activate extracellularly regulated mitogen-activated protein kinases ( P27361 /2 ) in normal and immortalized pituitary gonadotrophs and transfected cells expressing the P30968 . In immortalized hypothalamic DB00644 neurons ( GT1-7 cells ) , which also express DB00644 receptors , DB00644 , epidermal growth factor ( P01133 ) , and phorbol 12-myristate 13-acetate ( PMA ) caused marked phosphorylation of P27361 /2 . This action of DB00644 and PMA , but not that of P01133 , was primarily dependent on activation of protein kinase C ( PKC ) , and the P27361 /2 responses to all three agents were abolished by the selective P01133 receptor kinase inhibitor , AG1478 . Consistent with this , both DB00644 and P01133 increased tyrosine phosphorylation of the P01133 receptor . DB00644 and PMA , but not P01133 , caused rapid phosphorylation of the proline-rich tyrosine kinase , Pyk2 , at DB00135 (402) . This was reduced by Ca(2+) chelation and inhibition of PKC , but not by AG1478 . DB00644 stimulation caused translocation of PKC alpha and -epsilon to the cell membrane and enhanced the association of Src with PKC alpha and PKC epsilon , Pyk2 , and the P01133 receptor . The Src inhibitor , Q99463 , the C-terminal Src kinase ( Csk ) , and dominant-negative Pyk2 attenuated P27361 /2 activation by DB00644 and PMA but not by P01133 . These findings indicate that Src and Pyk2 act upstream of the P01133 receptor to mediate its transactivation , which is essential for DB00644 -induced P27361 /2 phosphorylation in hypothalamic DB00644 neurons .