An analysis of Lsm protein complexes

• Main Author: Reijns, Martin
• Published: University of Edinburgh 2006
• Subjects: 572.8
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.661038

Recombinant yeast Lsm proteins were purified from <i>Escherichia coli </i>and tested for their ability to promote annealing of the U4 and U6 snRNAs and to unwind a DNA/RNA duplex resembling the 3’ stem-loop of U6 snRNA. For comparison, Hfq, the Sm-like protein from <i>E. coli</i>, and recombinant human Lsm complexes were used in the same <i>in vitro</i> assays. The results are consistent with their ability to promote RNA/RNA annealing and to modulate RNA secondary structure, which, <i>in vivo</i>, may also allow them to affect RNA/protein interactions. The function of <i>LSM4</i> was studied by overexpression and depletion of wild-type Lsm4p, and by expression of an Lsm4p C-terminal deletion mutant. Results suggest that Lsm4p affects cell morphology and that its C-terminus promotes efficient recruitment of Lsm1-7p to P-bodies and may promote P-body formation. <i>LSM5</i> was shown to be dispensable for cell visibility, and its depletion was shown to affect levels of U4, U6 and U4/U6 RNAs similar to effects in <i>Ism6Δ </i>and <i>Ism7Δ </i>strains. The involvement of the different domains of Lsm1p and Lsm8p in localisation of these proteins to the cytoplasm (to P-bodies under stress conditions) and nucleus respectively was investigated by creating (deletion) mutants and hybrids of various domains of these proteins. Results suggest that the N-termini of both proteins play a central role in targeting them to their respective cellular locations. The <i>in vitro</i> studies reveal that the RNA chaperone function of Sm-like proteins appears to have been conserved from bacteria to eukaryotes. Presumably, gene duplication and formation of hetereo-multimeric complexes in higher organisms has allowed functional diversification.