D. Evans et al., A complex containing CstF-64 and the SL2 snRNP connects mRNA 3 ' end formation and trans-splicing in C-elegans operons, GENE DEV, 15(19), 2001, pp. 2562-2571
Polycistronic pre-mRNAs from Caenorhabditis elegans are processed by 3' end
formation of the upstream mRNA and SL2-specffic trans-splicing of the down
stream mRNA. These processes usually occur within an -100-nucleotide region
and are mechanistically coupled. In this paper, we report a complex in C.
elegans extracts containing the 3' end formation protein CstF-64 and the SL
2 snRNP. This complex, immunoprecipitated,with alpha CstF-64 antibody, cont
ains SL2 RNA, but not SL1 RNA or other U snRNAs. Using mutational analysis
we have been able to uncouple SL2 snRNP function and identity. SL2 RNA with
a mutation in stem/loop III is functional in vivo as a trans-splice donor,
but fails to splice to SL2-accepting trans-splice sites, suggesting that i
t has lost its identity as an SL2 snRNP. Importantly, stem/loop III mutatio
ns prevent association of SL2 RNA with CstF-64. In contrast, a mutation in
stem II that inactivates the SL2 snRNP still permits complex formation with
CstF-64. Therefore, SL2 RNA stem/loop III is required for both SL2 identit
y and formation of a complex containing CstF-64, but not for trans-splicing
. These results provide a molecular framework for the coupling of 3' end fo
rmation and trans-splicing in the processing of polycistronic pre-mRNAs fro
m C. elegans operons.