Stability of a PKCI-1-related mRNA is controlled by the splicing factor ASF/SF2: a novel function for SR proteins.
Pre-mRNA splicing is a widely used regulatory mechanism for controlling gene expression, and a family of conserved proteins, SR proteins, participate in both constitutive and alternative splicing. Here we describe a novel function for the SR protein ASF/SF2. We used an embryonic chicken cDNA library to screen for differential mRNA expression in the chicken B-cell line DT40-ASF, expressing or not expressing ASF/SF2. Remarkably, out of 3 x 10(6) clones screened, only one, isolated several times independently, showed ASF/SF2-related differential expression. The isolated cDNA, referred to here as PKCI-r (for PKCI-related), is closely related to the protein kinase C interacting protein (PKCI-1) gene. Transcript levels were increased approximately sixfold in ASF/SF2-depleted cells compared with cells expressing ASF/SF2, indicating a negative role for the SR protein. Strikingly, inhibition of ASF/SF2 expression had no significant effect on PKCI-r splicing, or transcription, but markedly increased the half-life of PKCI-r mRNA (6.6-fold). Similarly, increased mRNA stability was also observed upon expression of exogenous PKCI-r mRNA in cells depleted of ASF/SF2. ASF/SF2 bound to a discrete region containing a purine-rich sequence in the 3' UTR of the PKCI-r transcript, and deletion of this region eliminated ASF/SF2-mediated regulation of transcript stability. Together these data indicate a novel, direct effect of ASF/SF2 on PKCI-r mRNA stability. Therefore, ASF/SF2, and perhaps other SR proteins, affects gene expression in vertebrate cells through regulation of mRNA stability as well as splicing.
Lemaire
Raphael
R
Boston University School of Medicine, The Arthritis Center, Boston, Massachusetts 02118, USA.
Prasad
Jayendra
J
Kashima
Tsuyoshi
T
Gustafson
Jennifer
J
Manley
James L
JL
Lafyatis
Robert
R
D020413:3' Untranslated Regions / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002642:Chick Embryo; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics*; D009687:Nuclear Proteins / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011687:Purines; D012326:RNA Splicing; D020871:RNA Stability; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D000068103:Serine-Arginine Splicing Factors
D020413:3' Untranslated Regions; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D011687:Purines; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D000068103:Serine-Arginine Splicing Factors; D011493:Protein Kinase C