SMNrp is an essential pre-mRNA splicing factor required for the formation of the mature spliceosome

SMNrp, also termed SPF30, has recently been identified in spliceosomes asse mbled in vitro. We have functionally characterized this protein and show th at it is an essential splicing factor. We show that SMNrp is a 17S U2 snRNP -associated protein that appears in the pre-spliceosome (complex A) and the mature spliceosome (complex B) during splicing. Immunodepletion of SMNrp f rom nuclear extract inhibits the first step of pre-mRNA splicing by prevent ing the formation of complex B. Re-addition of recombinant SMNrp to immunod epleted extract reconstitutes both spliceosome formation and splicing. Muta tions in two domains of SMNrp, although similarly deleterious for splicing, differed in their consequences on U2 snRNP binding, suggesting that SMNrp may also engage in interactions with splicing factors other than the U2 snR NP. In agreement with this, we present evidence for an additional interacti on between SMNrp and the [U4/U6.U5] tri-snRNP. A candidate that may mediate this interaction, namely the U4/U6-90 kDa protein, has been identified. We suggest that SMNrp, as a U2 snRNP-associated protein, facilitates the recr uitment of the [U4/U6.U5] tri-snRNP to the pre-spliceosome.