Systematic identification of Ctr9 regulome in ERα-positive breast cancer.

Hao Zeng,Li Lu,Paul Ahlquist,Wei Xu,Mark Horswill,Xuehua Zhong,Ngai Ting Chan

doi:10.1186/s12864-016-3248-3

Abstract

BackgroundWe had previously identified Ctr9, the key scaffold subunit of the human RNA polymerase II (RNAPII) associated factor complex (PAFc), as a key factor regulating a massive ERα target gene expression and ERα-positive breast cancer growth. Furthermore, we have shown that knockdown of Ctr9 reduces ERα protein stability and decreases the occupancy of ERα and RNAPII at a few ERα-target loci. However, it remains to be determined whether Ctr9 controls ERα-target gene expression by regulating the global chromatin occupancy of ERα and RNAPII in the presence of estrogen.ResultsIn this study, we determined the genome-wide ERα and RNAPII occupancy in response to estrogen treatment and/or Ctr9 knockdown by performing chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq). We found that loss of Ctr9 dramatically decreases the global occupancy of ERα and RNAPII, highlighting the significance of Ctr9 in regulating estrogen signaling in ERα-positive breast cancer cells. Combining this resource with previously published genomic data sets, we identified a unique subset of ERα and Ctr9 target genes, and further delineated the independent function of Ctr9 from other subunits in PAFc when regulating transcription.ConclusionsOur data demonstrated that Ctr9, independent of other PAFc subunits, controls ERα-target gene expression by regulating global chromatin occupancies of ERα and RNAPII.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3248-3) contains supplementary material, which is available to authorized users.

Highlights

We had previously identified Ctr9, the key scaffold subunit of the human RNA polymerase II (RNAPII) associated factor complex (PAFc), as a key factor regulating a massive Estrogen receptor α (ERα) target gene expression and ERα-positive breast cancer growth
Ctr9 knockdown triggers genome-wide decrease of ERα occupancy To investigate the estrogen and Ctr9 mediated transcription regulation in ERα-positive breast cancer cells, we used the MCF7-tet-on-shCtr9 inducible knockdown cell line previously developed in our lab as a model system (Fig. 1a) [16, 33]
Consistent with our previous chromatin immunoprecipitation (ChIP)-qPCR results [16], we found that Ctr9 KD by Dox treatment significantly reduced ERα binding at the enhancer and promoter regions of TFF1 and GREB1 genes (Fig. 1b and c)

Summary

Introduction

We had previously identified Ctr, the key scaffold subunit of the human RNA polymerase II (RNAPII) associated factor complex (PAFc), as a key factor regulating a massive ERα target gene expression and ERα-positive breast cancer growth. We have shown that knockdown of Ctr reduces ERα protein stability and decreases the occupancy of ERα and RNAPII at a few ERα-target loci. It remains to be determined whether Ctr controls ERα-target gene expression by regulating the global chromatin occupancy of ERα and RNAPII in the presence of estrogen. Human RNA polymerase II (RNAPII) associated factor complex (PAFc) is composed of five subunits including PAF1, Ctr, Cdc ( known as parafibromin), Leo, and eukaryotic specific Ski ( known as WDR61) [5]. The PAFc regulated transcriptional elongation is coupled with histone modifications such as histone H2B mono-ubiquitination (H2Bub1), H3K4 trimethylation (H3K4me3), and H3K36 tri-methylation (H3K36me3) via protein-protein interaction of PAFc

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Discussion

Conclusion