A 69-KD PROTEIN THAT ASSOCIATES REVERSIBLY WITH THE SM CORE DOMAIN OFSEVERAL SPLICEOSOMAL SNRNP SPECIES

The biogenesis of the spliceosomal small nuclear ribonucleoproteins (s nRNPs) U1, U2, U4, and U5 involves: (a) migration of the snRNA molecul es from the nucleus to the cytoplasm; (b) assembly of a group of commo n proteins (Sm proteins) and their binding to a region on the snRNAs c alled the Sm-binding site; and (c) translocation of the RNP back to th e nucleus. A first prerequisite for understanding the assembly pathway and nuclear transport of the snRNPs in more detail is the knowledge o f all the snRNP proteins that play essential roles in these processes. We have recently observed a previously undetected 69-kD protein in 12 S U1 snRNPs isolated from HeLa nuclear extracts under non-denaturing c onditions that is clearly distinct from the U1-70K protein. The follow ing evidence indicates that the 69-kD protein is a common, rather than a U1-specific, protein, possibly associating with the snRNP core part icles by protein-protein interaction. (a) Antibodies raised against th e 69-kD protein, which did not cross-react with any of the Sm proteins B'-G, precipitated not only U1 snRNPs, but also the other spliceosoma l snRNPs U2, U4/U6 and U5, albeit to a lower extent. (b) U1, U2, and U 5 core RNP particles reconstituted in vitro contain the 69-kD protein. (c) Xenopus laevis oocytes contain an immunologically related homolog ue of the human 69-kD protein. When U1 snRNA as well as a mutant U1 sn RNA, that can bind the Sm core proteins but lacks the capacity to bind the U1-specific proteins 70K, A, and C, were injected into Xenopus oo cytes to allow assembly in vivo, they were recognized by antibodies sp ecific against the 69-kD protein in the ooplasm and in the nucleus. Th e 69-kD protein is under-represented, if present at all, in purified 1 7S U2 and in 25S [U4/U6.U5] tri-snRNPs, isolated from HeLa nuclear ext racts. Our results are consistent with the working hypothesis that thi s protein may either play a role in the cytoplasmic assembly of the co re domain of the snRNPs and/or in the nuclear transport of the snRNPs. After transport of the snRNPs into the nucleus, it may dissociate fro m the particles as for example in the case of the 17S U2 or the 25S [U 4/U6.U5] tri-snRNP, which bind more than 10 different snRNP specific p roteins each in the nucleus.