Cloning muscle isoforms of neural cell adhesion molecule using an episomal shuttle vector

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Abstract
Three distinct transcripts encoding two phosphatidylinositol (PI) linked isoforms of the neural cell adhesion molecule (NCAM) are induced during the differentiation of C2C12 myoblasts into myotubes. Corresponding NCAM clones were isolated from a mouse muscle cDNA library made in an Epstein-Barr virus shuttle vector that replicates extrachromosomally in human cells. Following transfection with the library, human cells expressing mouse NCAM were enriched using the fluorescence-activated cell sorter. Episomal NCAM clones recovered from sorted cells contain an 18-bp insert between exons 12 and 13. Two other NCAM cDNAs encode identical polypeptides containing a 108-bp insert homologous to the complete MSD1 domain, but differ in their 3′ untranslated regions. Induction of MSD1-containing transcripts in advance of myotube formation suggests that muscle-specific NCAMs contribute to myogenesis from the earliest stages of differentiation. Moreover, our studies demonstrate the feasibility of cloning tissue-specific molecules by transfection and expression of cDNA libraries in episomal vectors.