Two helices plus a linker: a small model substrate for eukaryotic RNase P.

Using precursor tRNA molecules to study RNA-protein interactions, we have identified an RNA motif recognized by eukaryotic RNase P (EC 3.1.26.5). Analysis of circularly permuted precursors indicates that interruptions in the sugar-phosphate backbone are not tolerated in the acceptor stem, in the T stem-loop, or between residues A-9 and G-10. Prokaryotic RNase P will function with a minihelix consisting of the acceptor stem connected directly to the T stem-loop. Eukaryotic RNase P cannot use such a minimal substrate unless a linker sequence is added in the gap where the D stem and anticodon stem-loop were deleted.

Distinct functions of SR proteins in recruitment of U1 small nuclear ribonucleoprotein to alternative 5' splice sites.

Alternative splicing of precursor messenger RNAs (pre-mRNAs) is an important mechanism for the regulation of gene expression. The members of the SR protein family of pre-mRNA splicing factors have distinct functions in promoting alternative splice site usage. Here we show that SR proteins are required for the first step of spliceosome assembly, interaction of the U1 small nuclear ribonucleoprotein complex (U1 snRNP) with the 5' splice site of the pre-mRNA. Further, we find that individual SR proteins have distinct abilities to promote interaction of U1 snRNP with alternative 5' splice junctions. These results suggest that SR proteins direct 5' splice site selection by regulation of U1 snRNP assembly onto the pre-mRNA.