Characterization of novel SF3b and 17S U2 snRNP proteins, including a human Prp5p homologue and an SF3b DEAD-box protein.
Mass spectrometry was used to identify novel proteins associated with the human 17S U2 snRNP and one of its stable subunits, SF3b. Several additional proteins were identified, demonstrating that 17S U2 snRNPs are significantly more complex than previously thought. Two of the newly identified proteins, namely the DEAD-box proteins SF3b125 and hPrp5 (a homologue of Saccharomyces cerevisiae Prp5p) were characterized further. Immunodepletion experiments with HeLa nuclear extract indicated that hPrp5p plays an important role in pre-mRNA splicing, acting during or prior to prespliceosome assembly. The SF3b-associated protein SF3b125 dissociates at the time of 17S U2 formation, raising the interesting possibility that it might facilitate the assembly of the 17S U2 snRNP. Finally, immunofluorescence/FISH studies revealed a differential subnuclear distribution of U2 snRNA, hPrp5p and SF3b125, which were enriched in Cajal bodies, versus SF3b155 and SF3a120, which were not; a model for 17S U2 snRNP assembly based on these findings is presented. Taken together, these studies provide new insight into the composition of the 17S U2 snRNP and the potential function of several of its proteins.
Will
Cindy L
CL
Department of Cellular Biochemistry, Max Planck Institute of Biophysical Chemistry, D-37077 Göttingen and EMBL, Bioanalytical Research Group, D-69117 Heidelberg, Germany.
Urlaub
Henning
H
Achsel
Tilmann
T
Gentzel
Marc
M
Wilm
Matthias
M
Lührmann
Reinhard
R
D053487:DEAD-box RNA Helicases; D006367:HeLa Cells; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D021122:Protein Subunits; D020365:RNA Helicases / Q000235:genetics / Q000378:metabolism*; D012326:RNA Splicing; D016601:RNA-Binding Proteins; D017413:Ribonucleoprotein, U2 Small Nuclear / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017381:Spliceosomes / Q000378:metabolism
D009687:Nuclear Proteins; D021122:Protein Subunits; D016601:RNA-Binding Proteins; D017413:Ribonucleoprotein, U2 Small Nuclear; C408625:DDX42 protein, human; C467585:DDX46 protein, human; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases