Inactivation of the SR protein splicing factor ASF/SF2 results in genomic instability.
SR proteins constitute a family of pre-mRNA splicing factors now thought to play several roles in mRNA metabolism in metazoan cells. Here we provide evidence that a prototypical SR protein, ASF/SF2, is unexpectedly required for maintenance of genomic stability. We first show that in vivo depletion of ASF/SF2 results in a hypermutation phenotype likely due to DNA rearrangements, reflected in the rapid appearance of DNA double-strand breaks and high-molecular-weight DNA fragments. Analysis of DNA from ASF/SF2-depleted cells revealed that the nontemplate strand of a transcribed gene was single stranded due to formation of an RNA:DNA hybrid, R loop structure. Stable overexpression of RNase H suppressed the DNA-fragmentation and hypermutation phenotypes. Indicative of a direct role, ASF/SF2 prevented R loop formation in a reconstituted in vitro transcription reaction. Our results support a model by which recruitment of ASF/SF2 to nascent transcripts by RNA polymerase II prevents formation of mutagenic R loop structures.
Li
Xialu
X
Department of Biological Sciences, Columbia University, New York, New York 10027, USA.
Manley
James L
JL
D000199:Actins / Q000235:genetics; D017398:Alternative Splicing / Q000235:genetics / Q000502:physiology; D000818:Animals; D016923:Cell Death / Q000502:physiology; D002460:Cell Line; D042822:Genomic Instability; D006801:Humans; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism*; D010641:Phenotype; D012319:RNA Polymerase II / Q000235:genetics; D012326:RNA Splicing / Q000235:genetics / Q000502:physiology; D016601:RNA-Binding Proteins; D016914:Ribonuclease H / Q000235:genetics / Q000378:metabolism; D000068103:Serine-Arginine Splicing Factors; D013997:Time Factors
D000199:Actins; D009687:Nuclear Proteins; D016601:RNA-Binding Proteins; D000068103:Serine-Arginine Splicing Factors; D012319:RNA Polymerase II; D016914:Ribonuclease H