Multiple cis-acting sequence elements are required for efficient splicing of simian virus 40 small-t antigen pre-mRNA.
We have determined the effects of a number of mutations in the small-t antigen mRNA intron on the alternative splicing pattern of the simian virus 40 early transcript. Expansion of the distance separating the small-t pre-mRNA lariat branch point and the shared large T-small t 3' splice site from 18 to 29 nucleotides (nt) resulted in a relative enhancement of small-t splicing in vivo. This finding, coupled with the observation that large-T pre-RNA splicing in vitro was not affected by this expansion, suggests that small-t splicing is specifically constrained by a short branch point-3' splice site distance. Similarly, the distance separating the 5' splice site and branch point (48 nt) was found to be at or near a minimum for small-t splicing, because deletions in this region as small as 2 nt dramatically reduced the ratio of small-t to large-T mRNA that accumulated in transfected cells. Finally, a specific sequence within the small-t intron, encompassing the upstream branch sites used in large-T splicing, was found to be an important element in the cell-specific pattern of early alternative splicing. Substitutions within this region reduced the ratio of small-t to large-T mRNA produced in HeLa cells but had only minor effects in human 293 cells.
Fu
X Y
XY
Department of Biological Sciences, Columbia University, New York, New York 10027.
Colgan
J D
JD
Manley
J L
JL
D000952:Antigens, Polyomavirus Transforming / Q000235:genetics*; D001483:Base Sequence; D002460:Cell Line; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D007438:Introns; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids; D012322:RNA Precursors / Q000235:genetics*; D012326:RNA Splicing; D013539:Simian virus 40 / Q000235:genetics*; D014158:Transcription, Genetic; D014162:Transfection
D000952:Antigens, Polyomavirus Transforming; D012322:RNA Precursors