Alternative splicing of the adenylyl cyclase stimulatory G-protein G alpha(s) is regulated by SF2/ASF and heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1) and involves the use of an unusual TG 3'-splice Site.
The factors involved in regulating alternative splicing of the human adenylyl cyclase stimulatory G-protein G alpha(s) in different cell types remain undefined. We have designed a G alpha(s) minigene that retains the signals required for G alpha(s) alternative splicing in vivo. Employing transient transfection of human myometrial smooth muscle cells and HeLa cells, as well as in vitro splicing assays, we have provided evidence that the antagonistic splicing factors SF2/ASF and hnRNPA1 act as potent regulators of G alpha(s) isoform expression in these cells. Both SF2/ASF and hnRNPA1 control the selection of competing 5'-splice sites and respectively promote inclusion or skipping of the small cassette-type exon 3 of G alpha(s) transcripts, resulting in the generation of G alpha(s)-long and G alpha(s)-short mRNA isoforms. We have also provided evidence that SF2/ASF and hnRNPA1 play a role in 3'-splice site selection involving the use of a non-canonical TG 3'-splice site preceding exon 4. Using a score-matrix analysis to identify putative exonic enhancer sequences (ESEs), we found multiple high score ESE motifs for SF2/ASF, SC35, and SRp40 in exon 3 of G alpha(s). These results suggest that tissue-specific expression of SF2/ASF and hnRNPA1 governs the expression of alternative isoforms of G alpha(s) in these different cells types.
Pollard
Alison J
AJ
Department of Obstetrics and Gynaecology, University of Newcastle upon Tyne, Royal Victoria Infirmary, Richardson Road, Newcastle upon Tyne, NE1 4LP, United Kingdom. A.J.Pollard@ncl.ac.uk
Krainer
Adrian R
AR
Robson
Stephen C
SC
Europe-Finner
G Nicholas
GN
D000262:Adenylyl Cyclases / Q000378:metabolism*; D017398:Alternative Splicing; D001483:Base Sequence; D017931:DNA Primers; D005260:Female; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000235:genetics*; D005786:Gene Expression Regulation; D006367:HeLa Cells; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D008969:Molecular Sequence Data; D009130:Muscle, Smooth / Q000378:metabolism; D009215:Myometrium / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms / Q000235:genetics; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics; D016601:RNA-Binding Proteins; D015183:Restriction Mapping; D012261:Ribonucleoproteins / Q000378:metabolism*; D000068103:Serine-Arginine Splicing Factors; D014158:Transcription, Genetic
D017931:DNA Primers; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D009687:Nuclear Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; C000619656:hnRNPA1 protein, human; D000068103:Serine-Arginine Splicing Factors; D019205:GTP-Binding Protein alpha Subunits, Gs; D000262:Adenylyl Cyclases