A conserved epitope on a subset of SR proteins defines a larger family of Pre-mRNA splicing factors.
The removal of introns from eukaryotic pre-mRNA occurs in a large ribonucleoprotein complex called the spliceosome. We have generated a monoclonal antibody (mAb 16H3) against four of the family of six SR proteins, known regulators of splice site selection and spliceosome assembly. In addition to the reactive SR proteins, SRp20, SRp40, SRp55, and SRp75, mAb 16H3 also binds approximately 20 distinct nuclear proteins in human, frog, and Drosophila extracts, whereas yeast do not detectably express the epitope. The antigens are shown to be nuclear, nonnucleolar, and concentrated at active sites of RNA polymerase II transcription which suggests their involvement in pre-mRNA processing. Indeed, most of the reactive proteins observed in nuclear extract are detected in spliceosomes (E and/or B complex) assembled in vitro, including the U1 70K component of the U1 small nuclear ribonucleoprotein particle and both subunits of U2AF. Interestingly, the 16H3 epitope was mapped to a 40-amino acid polypeptide composed almost exclusively of arginine alternating with glutamate and aspartate. All of the identified antigens, including the human homolog of yeast Prp22 (HRH1), contain a similar structural element characterized by arginine alternating with serine, glutamate, and/or aspartate. These results indicate that many more spliceosomal components contain such arginine-rich domains. Because it is conserved among metazoans, we propose that the "alternating arginine" domain recognized by mAb 16H3 may represent a common functional element of pre-mRNA splicing factors.
Neugebauer
K M
KM
Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, Washington 98104, USA.
Stolk
J A
JA
Roth
M B
MB
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D002451:Cell Compartmentation; D004330:Drosophila; D000939:Epitopes; D006801:Humans; D007150:Immunohistochemistry; D007739:L Cells; D051379:Mice; D009687:Nuclear Proteins / Q000276:immunology* / Q000302:isolation & purification; D012322:RNA Precursors / Q000378:metabolism; D012326:RNA Splicing / Q000276:immunology*; D016601:RNA-Binding Proteins / Q000145:classification / Q000276:immunology* / Q000302:isolation & purification; D011898:Ranidae; D017412:Ribonucleoprotein, U1 Small Nuclear; D017381:Spliceosomes / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D009687:Nuclear Proteins; D012322:RNA Precursors; D016601:RNA-Binding Proteins; D017412:Ribonucleoprotein, U1 Small Nuclear