Regulation of Heat Shock Gene Expression by RNA Polymerase II Elongation Factor, Elongin A

Abstract

The elongation stage of transcription by RNA polymerase II (Pol II) has emerged as an essential regulated step. Elongin A (EloA) is the largest subunit of the Elongin complex that can increase the catalytic rate of mRNA synthesis by Pol II. We recently demonstrated that the Elongin A homologue in Drosophila, dEloA, is essential and has properties consistent with those of a Pol II elongation factor in vivo. The goal of this study was to test whether dEloA is required for heat shock gene transcription, since heat shock gene expression is thought to be controlled at the level of Pol II elongation. Here, we demonstrate that dEloA is rapidly recruited to heat shock loci with Pol II in response to heat shock. Furthermore, through the use of RNA interference in vivo, we show that dEloA is required for the proper expression of one of these genes, HSP70, and that its requirement for heat shock gene expression is exerted after the initiation of transcription at heat shock loci. Our data represent the first demonstration of an essential role for an RNA polymerase II elongation factor in the regulation of heat shock gene expression in an animal model.The Elongin complex was originally purified based on its ability to stimulate Pol II 1 elongation in an in vitro transcription elongation assay (1). Elongin is a heterotrimeric complex, consisting of A, B, and C subunits, which are ϳ110, 18, and 15 kilodaltons in size, respectively. Elongin A is the transcriptionally active subunit of the complex, and the addition of this protein alone to in vitro transcription reactions is sufficient to stimulate transcription elongation by Pol II (1-3). In the absence of Elongin A, Elongin BC is unable to enhance Pol II processivity.A number of Elongin A family members have been identified. They include three mammalian homologues (Elongin A, A2, and A3) and an Elongin A homologue in Caenorhabditis elegans (1,4,5). Each of these proteins is capable of stimulating Pol II elongation in vitro and interacting with the Elongin BC complex.Recently, we identified and characterized the Drosophila melanogaster homologue of Elongin A, named dEloA (14). Like the other Elongin A family members, dEloA stimulates Pol II elongation activity in vitro. Additionally, dEloA is concentrated at transcriptionally active loci on Drosophila polytene chromosomes, and it colocalizes extensively with transcriptionally active isoforms of Pol II. Consistent with this observation, dEloA interacts with Pol II in Drosophila extracts. Collectively, these observations are consistent with the hypothesis that the Elongin proteins are Pol II-associated factors in vivo.Heat shock gene expression is thought to be controlled at the level of promoter clearance (16). Therefore, the expression of heat shock genes should be particularly sensitive to the cellular level of Pol II elongation factors. A variety of Pol II elongation factors are recruited to sites of heat shock gene expression during heat shock (6 -9, 15). However, very little is known about the role(s) of these factors in th...

The super elongation complex (SEC) and MLL in development and disease

“…Paused RNA Pol II has been best characterized as a regulatory point at heat-shock loci in Drosophila, where RNA Pol II sits in an engaged state at the promoter, nonproductively transcribing ;25 nucleotides (nt) in the nonstressed state. Upon heat shock, Pol II is released for the production of full-length transcripts (Gilmour and Lis 1986;Rougvie and Lis 1988;Gerber et al 2001Gerber et al , 2005Smith et al 2008b). …”

Transcriptional blackjack with p21: Figure 1.

“…Loss of CDK9 activity is followed by a substantial decrease in the association of other essential Pol II elongation factors such as FACT; however, p21 gene expression is not altered in their absence. Recently, several RNA Pol II elongation factors, such as ELL and Elongin A, were found to be required for proper cellular response to heat shock (Gerber et al 2005). It will be interesting to uncover whether such Pol II elongation factors that regulate the elongation stage of transcription are required for proper p21 gene expression.…”

Induction of apoptosis and cellular senescence in mice lacking transcription elongation factor, Elongin A

“…8,13,14 Although the functions of Elongin A in vivo remain largely unclear, Gerber et al 15 have recently reported that the Drosophila homolog of Elongin A (dEloA) colocalizes with pol II at sites of active transcription on polytene chromosomes, and it also plays a critical role in heat shock gene expression in vivo. 15,16 Moreover, by the RNA interference (RNAi) knockdown of dEloA, they have also shown it to be an essential factor required for proper metamorphosis, although the relationship between the elongation activity and viability remains unclear.To gain more insight into the physiological functions of Elongin A, we have generated Elongin A-deficient mice. Elongin A À/À embryos exhibited severe growth retardation and died at between days 10.5 and 12.5 of gestation, most likely owing to excessive apoptosis.…”