Distinct NCAM splicing events are differentially regulated during rat brain development

Abstract

Primary transcripts for the neural cell adhesion molecule NCAM are highly alternatively spliced potentially giving rise to over 100 different mRNA forms. These mRNAs encode three major polypeptide isoforms of 120, 140, and 180 kDa each of which is thought to be compsed of a mixture of polypeptides that differ by the variable presence of small exons at two locations. These NCAM ‘microsplicing’ patterns were examined within mRNA populations encoding each of the major isoforms to identify exactly which NCAM forms are present during brain development. The proportion of NCAM mRNAs containing at the exon 7 8 junction the alternatively microspliced 30 bp VASE exon increased similarly during brain development in mRNAs encoding all three major size classes. Perinatal brain, cultures of neurons from embryonic rats, and of glia from newborn rats all had low VASE levels while about 50% of the mRNAs in adult brain expressed VASE. In contrast, microsplicing at the exon 12/exon 13 junction was differentially regulated among NCAM major size classes. mRNAs containing microspliced exons totaling 3, 15, or 18 base pairs (bp) represented greater than 50% of the total mRNAs encoding the 120 kDa forms at all ages. However, these exons were present in less than 15% of the 140 and 180 kDa encoding mRNAs in rats older than embryonic day 15. Similar results at the exon 12 13 junction were observed with mRNAs from neuronal cultures while glial cultures had greater levels of a 3 bp pair exon at this junction. Analyses of both primary and clonal neuronal and glial cell cultures indicate that cell phenotype is not the dominant factor in regulating NCAM microsplicing of VASE but may be a more significant factor at the exon 12 13 junction. Comparison to true immunoglobulin suggest that alternative splicing at the exon 12 13 junction could be placed a ‘bend’ in the NCAM polypeptide. Thus, most glial NCAM could have a distinct three-dimensional configuration compared to neuronal NCAM. In conclusion, these experiments have determined the relative abundance of microspliced NCAM mRNA forms during neural development and clearly identified differences in alternative microsplicing among mRNAs encoding the major NCAM isoforms.