Establishment of Centromeric Chromatin by the CENP-A Assembly Factor CAL1 Requires FACT-Mediated Transcription

Abstract

Centromeres are essential chromosomal structures that mediate accurate chromosome segregation during cell division. Centromeres are specified epigenetically by the heritable incorporation of the centromeric histone H3 variant CENP-A. While many of the primary factors that mediate centromeric deposition of CENP-A are known, the chromatin and DNA requirements of this process have remained elusive. Here, we uncover a role for transcription in Drosophila CENP-A deposition. Using an inducible ectopic centromere system that uncouples CENP-A deposition from endogenous centromere function and cell-cycle progression, we demonstrate that CENP-A assembly by its loading factor, CAL1, requires RNAPII-mediated transcription of the underlying DNA. This transcription depends on the CAL1 binding partner FACT, but not on CENP-A incorporation. Our work establishes RNAPII passage as a key step in chaperone-mediated CENP-A chromatin establishment and propagation.

Additional Information

© 2015 Elsevier. Under an Elsevier user license. Received 7 October 2014, Revised 9 April 2015, Accepted 18 May 2015, Available online 6 July 2015. We thank Bo Reese (CGI) for sequencing support, G. Karpen, S. Hirose, A. Straight, and P. Heun for reagents, I. Cheeseman for critically reading the manuscript, L. Core for suggestions and help with ChIP-chip data analysis, and the Drosophila RNAi Screening Center (DRSC) for resources. This work was supported by awards from the NSF (1024973) (to B.G.M.) and the NIH (GM108829) (to B.G.M.). D.M.G. is supported by grants from Cancer Research UK (C3/A11431) and the Medical Research Council (G1001696); Z.L. is supported by a long-term FEBS Fellowship; and R.J.O. is supported by an NSF award (1244146). Accession Numbers. The accession number for the ChIP-seq and RNA-seq raw data reported in this paper is NCBI: SRP059507.

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Supplemental Material - 1-s2.0-S1534580715003482-mmc1.pdf

Accepted Version - nihms693741.pdf

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