The U1 snRNP-associated factor Luc7p affects 5' splice site selection in yeast and human.
yLuc7p is an essential subunit of the yeast U1 snRNP and contains two putative zinc fingers. Using RNA-protein cross-linking and directed site-specific proteolysis (DSSP), we have established that the N-terminal zinc finger of yLuc7p contacts the pre-mRNA in the 5' exon in a region close to the cap. Modifying the pre-mRNA sequence in the region contacted by yLuc7p affects splicing in a yLuc7p-dependent manner indicating that yLuc7p stabilizes U1 snRNP-pre-mRNA interaction, thus reminding of the mode of action of another U1 snRNP component, Nam8p. Database searches identified three putative human yLuc7p homologs (hLuc7A, hLuc7B1 and hLuc7B2). These proteins have an extended C-terminal tail rich in RS and RE residues, a feature characteristic of splicing factors. Consistent with a role in pre-mRNA splicing, hLuc7A localizes in the nucleus and antibodies raised against hLuc7A specifically co-precipitate U1 snRNA from human cell extracts. Interestingly, hLuc7A overexpression affects splicing of a reporter in vivo. Taken together, our data suggest that the formation of a wide network of protein-RNA interactions around the 5' splice site by U1 snRNP-associated factors contributes to alternative splicing regulation.
Puig
Oscar
O
European Molecular Biology Laboratory, Meyerhofstrasse, 1, 69117 Heidelberg, Germany. oscar.puig@helsinki.fi
Bragado-Nilsson
Elisabeth
E
Koski
Terhi
T
Séraphin
Bertrand
B
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D001665:Binding Sites; D005091:Exons; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D012322:RNA Precursors / Q000737:chemistry / Q000378:metabolism; D022821:RNA Splice Sites; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000302:isolation & purification; D016335:Zinc Fingers
D005656:Fungal Proteins; C404083:LUC7L3 protein, human; D009687:Nuclear Proteins; D012322:RNA Precursors; D022821:RNA Splice Sites; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D017412:Ribonucleoprotein, U1 Small Nuclear