Nucleic Acids Research, 2002, Vol. 30, No. 15 3360-3367
© 2002 Oxford University Press
Analysis of the role of Caenorhabditis elegans GC-AG introns in regulated splicing
Department of MCD Biology and Center for Molecular Biology of RNA, Sinsheimer Laboratories, University of California, Santa Cruz, CA 95064, USA
*To whom correspondence should be addressed. Tel: +1 831 459 5131; Fax: +1 831 459 3737; Email: zahler{at}biology.ucsc.edu
Present addresses:
A. Brock Roller, University of Rochester School of Medicine, Rochester, NY 14642, USA
W. James Kent, UCSC Center for Genomic Sciences, University of California, Santa Cruz, Santa Cruz, CA 95064, USA
GC-AG introns represent 0.7% of total human pre-mRNA introns. To study the function of GC-AG introns in splicing regulation, 196 cDNA-confirmed GC-AG introns were identified in Caenorhabditis elegans. These represent 0.6% of the cDNA- confirmed intron data set for this organism. Eleven of these GC-AG introns are involved in alternative splicing. In a comparison of the genomic sequences of homologous genes between C.elegans and Caenorhabditis briggsae for 26 GC-AG introns, the C at the +2 position is conserved in only five of these introns. A system to experimentally test the function of GC-AG introns in alternative splicing was developed. Results from these experiments indicate that the conserved C at the +2 position of the tenth intron of the let-2 gene is essential for developmentally regulated alternative splicing. This C allows the splice donor to function as a very weak splice site that works in balance with an alternative GT splice donor. A weak GT splice donor can functionally replace the GC splice donor and allow for splicing regulation. These results indicate that while the majority of GC-AG introns appear to be constitutively spliced and have no evolutionary constraints to prevent them from being GT-AG introns, a subset of GC-AG introns is involved in alternative splicing and the C at the +2 position of these introns can have an important role in splicing regulation.
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