Histone H3.3 Is Required to Maintain Replication Fork Progression after UV Damage

• Published in: Current biology Vol. 24; no. 18; pp. 2195 - 2201
• Main Authors: Frey, Alexander, Listovsky, Tamar, Guilbaud, Guillaume, Sarkies, Peter, Sale, Julian E.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 22.09.2014; Cell Press
• Subjects: Animals; Avian Proteins - metabolism; Cell Line, Tumor; Chickens; Chromatin - metabolism; DNA Damage; DNA Repair - radiation effects; DNA Replication - radiation effects; Histones - metabolism; Ultraviolet Rays - adverse effects
• ISSN: 0960-9822; 1879-0445
• DOI: 10.1016/j.cub.2014.07.077

Unlike histone H3, which is present only in S phase, the variant histone H3.3 is expressed throughout the cell cycle [1] and is incorporated into chromatin independent of replication [2]. Recently, H3.3 has been implicated in the cellular response to ultraviolet (UV) light [3]. Here, we show that chicken DT40 cells completely lacking H3.3 are hypersensitive to UV light, a defect that epistasis analysis suggests may result from less-effective nucleotide excision repair. Unexpectedly, H3.3-deficient cells also exhibit a substantial defect in maintaining replication fork progression on UV-damaged DNA, which is independent of nucleotide excision repair, demonstrating a clear requirement for H3.3 during S phase. Both the UV hypersensitivity and replication fork slowing are reversed by expression of H3.3 and require the specific residues in the α2 helix that are responsible for H3.3 binding its dedicated chaperones. However, expression of an H3.3 mutant in which serine 31 is replaced with alanine, the equivalent residue in H3.2, restores normal fork progression but not UV resistance, suggesting that H3.3[S31A] may be incorporated at UV-damaged forks but is unable to help cells tolerate UV lesions. Similar behavior was observed with expression of H3.3 carrying mutations at K27 and G34, which have been reported in pediatric brain cancers. We speculate that incorporation of H3.3 during replication may mark sites of lesion bypass and, possibly through an as-yet-unidentified function of the N-terminal tail, facilitate subsequent processing of the damage. •We report a vertebrate cell line completely lacking the histone variant H3.3•H3.3-deficient cells are hypersensitive to DNA damage•A supply of H3.3 is required to maintain fork progression after UV damage•This S phase role requires the distinct chaperone-binding patch of H3.3 Frey et al. describe a vertebrate cell line completely lacking histone variant H3.3 and show that a supply of this histone variant is required to maintain processive replication after exposure to UV light and cellular resistance to DNA damage.