Journal
GENES & DEVELOPMENT
Volume 28, Issue 7, Pages 735-748Publisher
COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT
DOI: 10.1101/gad.234070.113
Keywords
FACT; transcription-replication collisions; R loops; genome instability; chromatin reorganization
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Funding
- Spanish Ministry of Economy and Competitiveness [Consolider CSD2007-00015, BFU2010-16372]
- Junta de Andalucia [CVI4567]
- European Union (FEDER)
- Spanish Ministry of Health
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FACT (facilitates chromatin transcription) is a chromatin-reorganizing complex that swaps nucleosomes around the RNA polymerase during transcription elongation and has a role in replication that is not fully understood yet. Here we show that recombination factors are required for the survival of yeast FACT mutants, consistent with an accumulation of DNA breaks that we detected by Rad52 foci and transcription-dependent hyperrecombination. Breaks also accumulate in FACT-depleted human cells, as shown by gamma H2AX foci and single-cell electrophoresis. Furthermore, FACT-deficient yeast and human cells show replication impairment, which in yeast we demonstrate by ChIP-chip (chromatin immunoprecipitation [ChIP] coupled with microarray analysis) of Rrm3 to occur genome-wide but preferentially at highly transcribed regions. Strikingly, in yeast FACT mutants, high levels of Rad52 foci are suppressed by RNH1 overexpression; R loops accumulate at high levels, and replication becomes normal when global RNA synthesis is inhibited in FACT-depleted human cells. The results demonstrate a key function of FACT in the resolution of R-loop-mediated transcription-replication conflicts, likely associated with a specific chromatin organization.
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