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. 1998 Feb;4(2):127–141.

The 20kD protein of human [U4/U6.U5] tri-snRNPs is a novel cyclophilin that forms a complex with the U4/U6-specific 60kD and 90kD proteins.

S Teigelkamp 1, T Achsel 1, C Mundt 1, S F Göthel 1, U Cronshagen 1, W S Lane 1, M Marahiel 1, R Lührmann 1
PMCID: PMC1369602  PMID: 9570313

Abstract

Cyclophilins (Cyps) catalyze the cis/trans isomerization of peptidyl-prolyl bonds, a rate-limiting step in protein folding. In some cases, cyclophilins have also been shown to form stable complexes with specific proteins in vivo and may thus also act as chaperone-like molecules. We have characterized the 20kD protein of the spliceosomal 25S [U4/U6.U5] tri-snRNP complex from HeLa cells and show that it is a novel human cyclophilin (denoted SnuCyp-20). Purified [U4/U6.U5] tri-snRNPs, but not U1, U2, or U5 snRNPs, exhibit peptidyl-prolyl cis/trans isomerase activity in vitro, which is cyclosporin A-sensitive, suggesting that SnuCyp-20 is an active isomerase. Consistent with its specific association with tri-snRNPs in vitro, immunofluorescence microscopy studies showed that SnuCyp-20 is predominantly located in the nucleus, where it colocalizes in situ with typical snRNP-containing structures referred to as nuclear speckles. As a first step toward the identification of possible targets of SnuCyp-20, we have investigated the interaction of SnuCyp-20 with other proteins of the tri-snRNP. Fractionation of RNA-free protein complexes dissociated from isolated tri-snRNPs by treatment with high salt revealed that SnuCyp-20 is part of a biochemically stable heteromer containing additionally the U4/U6-specific 60kD and 90kD proteins. By coimmunoprecipitation experiments performed with in vitro-translated proteins, we could further demonstrate a direct interaction between SnuCyp-20 and the 60kD protein, but failed to detect a protein complex containing the 90kD protein. The formation of a stable SnuCyp-20/60kD/90kD heteromer may thus require additional factors not present in our in vitro reconstitution system. We discuss possible roles of SnuCyp-20 in the assembly of [U4/U6.U5] tri-snRNPs and/or in conformational changes occurring during the splicing process.

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