Affiliation:
1. Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York
Abstract
ABSTRACT
The uninduced
Drosophila hsp70
gene is poised for rapid activation. Here we examine the rapid changes upon heat shock in levels and location of heat shock factor (HSF), RNA polymerase II (Pol II) and its phosphorylated forms, and the Pol II kinase P-TEFb on
hsp70
in vivo by using both real-time PCR assays of chromatin immunoprecipitates and polytene chromosome immunofluorescence. These studies capture Pol II recruitment and progression along
hsp70
and reveal distinct spatial and temporal patterns of serine 2 and serine 5 phosphorylation: in uninduced cells, the promoter-paused Pol II shows Ser5 but not Ser2 phosphorylation, and in induced cells the relative level of Ser2-P Pol II is lower at the promoter than at regions downstream. An early time point of heat shock activation captures unphosphorylated Pol II recruited to the promoter prior to P-TEFb, and during the first wave of transcription Pol II and the P-TEFb kinase can be seen tracking together across
hsp70
with indistinguishable kinetics. Pol II distributions on several other genes with paused Pol II show a pattern of Ser5 and Ser2 phosphorylation similar to that of
hsp70
. These studies of factor choreography set important limits in modeling transcription regulatory mechanisms.
Publisher
American Society for Microbiology
Subject
Cell Biology,Molecular Biology
Cited by
196 articles.
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