In vivo evidence that defects in the transcriptional elongation factors RPB2, TFIIS, and SPT5 enhance upstream poly(A) site utilization.
While a number of proteins are involved in elongation processes, the mechanism for action of most of these factors remains unclear primarily because of the lack of suitable in vivo model systems. We identified in yeast several genes that contain internal poly(A) sites whose full-length mRNA formation is reduced by mutations in RNA polymerase II subunit RPB2, elongation factor SPT5, or TFIIS. RPB2 and SPT5 defects also promoted the utilization of upstream poly(A) sites for genes that contain multiple 3' poly(A) signaling sequences, supporting a role for elongation in differential poly(A) site choice. Our data suggest that elongation defects cause increased transcriptional pausing or arrest that results in increased utilization of internal or upstream poly(A) sites. Transcriptional pausing or arrest can therefore be visualized in vivo if a gene contains internal poly(A) sites, allowing biochemical and genetic study of the elongation process.
Cui
Yajun
Y
Department of Biochemistry and Molecular Biology, University of New Hampshire, Durham, New Hampshire 03824, USA.
Denis
Clyde L
CL
D000483:Alleles; D000818:Animals; D002868:Chromosomal Proteins, Non-Histone; D005800:Genes, Fungal; D017930:Genes, Reporter; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism*; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D035165:Transcription Factors, General / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors / Q000235:genetics / Q000378:metabolism*; D039221:mRNA Cleavage and Polyadenylation Factors / Q000378:metabolism
D002868:Chromosomal Proteins, Non-Histone; D009687:Nuclear Proteins; D021122:Protein Subunits; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D035165:Transcription Factors, General; D035602:Transcriptional Elongation Factors; D039221:mRNA Cleavage and Polyadenylation Factors; C018648:transcription factor S-II; C069940:SPT5 transcriptional elongation factor; D012319:RNA Polymerase II