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Forced Expression of MyoD Creates a Muscle-Specific Test System for Exogenous Genes

A new approach free of time-consuming cloning procedures is described using MRF4 regulatory sequences acting as a physiological and muscular-specific promoter on human tissues; this method is useful for testing the expression of exogenous genes in cultured myoblasts and myotubes. pMRF4Z is a plasmid containing the regulatory sequences of Herculin (or MRF4, or MyF6), a gene of the MyoD family, driving the expression of a reporter gene (E.Coli (3-galactosidase or "lacZ"). The DNAs were introduced into the cultured cells by the biolistic gun: mouse embryo-derived cells and human fibroblasts were bombarded with high velocity micro projectiles each carrying at the same time two different plasmids. The pair of plasmids were a MyoD construct together with pMRF4Z or pCMVlacZ, in the latter plasmid a viral promoter drives the expression of a reporter gene. The forced expression of MyoD family genes induced myogenesis and several days after the bombardment polynucleated myotubes were positive for lacZ activity.