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

• CENP-A deposition is coupled with transcription
• CAL1 recruits RNAPII onto DNA during CENP-A deposition
• CAL1 interacts directly with the histone chaperone FACT
• FACT depletion causes loss of transcription and defective CENP-A deposition

SummaryCentromeres 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.

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