A mammalian germ cell-specific RNA-binding protein interacts with ubiquitously expressed proteins involved in splice site selection.
RNA-binding motif (RBM) genes are found on all mammalian Y chromosomes and are implicated in spermatogenesis. Within human germ cells, RBM protein shows a similar nuclear distribution to components of the pre-mRNA splicing machinery. To address the function of RBM, we have used protein-protein interaction assays to test for possible physical interactions between these proteins. We find that RBM protein directly interacts with members of the SR family of splicing factors and, in addition, strongly interacts with itself. We have mapped the protein domains responsible for mediating these interactions and expressed the mouse RBM interaction region as a bacterial fusion protein. This fusion protein can pull-down several functionally active SR protein species from cell extracts. Depletion and add-back experiments indicate that these SR proteins are the only splicing factors bound by RBM which are required for the splicing of a panel of pre-mRNAs. Our results suggest that RBM protein is an evolutionarily conserved mammalian splicing regulator which operates as a germ cell-specific cofactor for more ubiquitously expressed pre-mRNA splicing activators.
Elliott
D J
DJ
Medical Research Council Human Genetics Unit, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, Scotland. davide@hgu.mrc.ac.uk
Bourgeois
C F
CF
Klink
A
A
Stévenin
J
J
Cooke
H J
HJ
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D005854:Germ Cells / Q000378:metabolism; D006801:Humans; D007248:Infertility, Male / Q000378:metabolism; D008297:Male; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism*; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012322:RNA Precursors / Q000378:metabolism; D012326:RNA Splicing; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D000068103:Serine-Arginine Splicing Factors; D013379:Substrate Specificity; D020798:Two-Hybrid System Techniques
D009687:Nuclear Proteins; D010446:Peptide Fragments; D010750:Phosphoproteins; C501499:RBMY1A1 protein, human; D012322:RNA Precursors; D016601:RNA-Binding Proteins; C501500:Rbmy protein, mouse; D011993:Recombinant Fusion Proteins; C491388:SRSF3 protein, human; C106161:Srsf3 protein, mouse; D000068103:Serine-Arginine Splicing Factors