Drosophila Paf1 modulates chromatin structure at actively transcribed genes.
The Paf1 complex in yeast has been reported to influence a multitude of steps in gene expression through interactions with RNA polymerase II (Pol II) and chromatin-modifying complexes; however, it is unclear which of these many activities are primary functions of Paf1 and are conserved in metazoans. We have identified and characterized the Drosophila homologs of three subunits of the yeast Paf1 complex and found striking differences between the yeast and Drosophila Paf1 complexes. We demonstrate that although Drosophila Paf1, Rtf1, and Cdc73 colocalize broadly with actively transcribing, phosphorylated Pol II, and all are recruited to activated heat shock genes with similar kinetics; Rtf1 does not appear to be a stable part of the Drosophila Paf1 complex. RNA interference (RNAi)-mediated depletion of Paf1 or Rtf1 leads to defects in induction of Hsp70 RNA, but tandem RNAi-chromatin immunoprecipitation assays show that loss of neither Paf1 nor Rtf1 alters the density or distribution of phosphorylated Pol II on the active Hsp70 gene. However, depletion of Paf1 reduces trimethylation of histone H3 at lysine 4 in the Hsp70 promoter region and significantly decreases the recruitment of chromatin-associated factors Spt6 and FACT, suggesting that Paf1 may manifest its effects on transcription through modulating chromatin structure.
Adelman
Karen
K
Department of Molecular Biology and Genetics, Biotechnology Building, Cornell University, Ithaca, NY 14853, USA.
Wei
Wenxiang
W
Ardehali
M Behfar
MB
Werner
Janis
J
Zhu
Bing
B
Reinberg
Danny
D
Lis
John T
JT
D000818:Animals; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation; D004257:DNA Polymerase II / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D029721:Drosophila Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000235:genetics* / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D006609:High Mobility Group Proteins / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D008745:Methylation; D010445:Peptide Elongation Factors / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D035602:Transcriptional Elongation Factors / Q000378:metabolism
D002843:Chromatin; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D018840:HSP70 Heat-Shock Proteins; D006609:High Mobility Group Proteins; D006657:Histones; C508978:Hyx protein, Drosophila; D010445:Peptide Elongation Factors; D012333:RNA, Messenger; C508977:Rtf1 protein, Drosophila; C489543:SSRP protein, Drosophila; C470423:Spt6 protein, Drosophila; D035602:Transcriptional Elongation Factors; C508976:atms protein, Drosophila; D004257:DNA Polymerase II; D008239:Lysine