During DNA replication, chromatin is reassembled by recycling of modified old histones and deposition of new ones. How histone dynamics integrates with DNA replication to maintain genome and epigenome information remains unclear. Here, we reveal how human MCM2, part of the replicative helicase, chaperones histones H3–H4. Our first structure shows an H3–H4 tetramer bound by two MCM2 histone-binding domains (HBDs), which hijack interaction sites used by nucleosomal DNA. Our second structure reveals MCM2 and ASF1 cochaperoning an H3–H4 dimer. Mutational analyses show that the MCM2 HBD is required for MCM2–7 histone-chaperone function and normal cell proliferation. Further, we show that MCM2 can chaperone both new and old canonical histones H3–H4 as well as H3.3 and CENPA variants. The unique histone-binding mode of MCM2 thus endows the replicative helicase with ideal properties for recycling histones genome wide during DNA replication.

Summary After DNA replication, chromosomal processes including DNA repair and transcription take place in the context of sister chromatids. While cell cycle regulation can guide these processes globally, mechanisms to distinguish pre- and post-replicative states locally remain unknown. Here, we reveal that new histones incorporated during DNA replication provide a signature of post-replicative chromatin, read by the TONSL–MMS22L1–4 homologous recombination (HR) complex. We identify the TONSL Ankyrin Repeat Domain (ARD) as a reader of histone H4 tails unmethylated at K20 (H4K20me0), which are specific to new histones incorporated during DNA replication and mark post-replicative chromatin until G2/M. Accordingly, TONSL–MMS22L binds new histones H3–H4 both prior to and after incorporation into nucleosomes, remaining on replicated chromatin until late G2/M. H4K20me0 recognition is required for TONSL–MMS22L binding to chromatin and accumulation at challenged replication forks and DNA lesions. Consequently, TONSL ARD mutants are toxic, compromising genome stability, cell viability and resistance to replication stress. Together, this reveals a histone reader based mechanism to recognize the post-replicative state, offering a new approach and opportunity to understand DNA repair with potential for targeted cancer therapy.

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TLKs promote nucleosome assembly and prevent fork collapse and DNA damage upon checkpoint or PARP inhibition.

Homologous recombination (HR) is an important route for repairing DNA double-strand breaks (DSBs). The early stages of HR are well understood, but later stages remain mysterious. In this issue of Genes & Development, Hustedt and colleagues (pp. 1397-1415) reveal HROB as a new player in HR required for recruitment of the MCM8-9 complex, which is paralogous to the MCM2-7 replicative helicase. HROB functions closely with MCM8-9 to promote postsynaptic DNA repair synthesis. This study sheds valuable light on late events in HR and suggests that HROB may load MCM8-9 onto HR intermediates to facilitate the DNA unwinding required for DNA repair synthesis.Homologous recombination (HR) is a major route for repairing DNA double-strand breaks (DSBs) (Prakash et al. 2015). HR starts with resection of broken DNA ends, involving nucleases and helicases. The resulting overhangs, coated by RAD51 to yield a nucleoprotein filament, invade the intact sister chromatid, searching for a homologous sequence. The pairing of RAD51-coated single-stranded DNA (ssDNA) with the donor strand (termed synapsis) results in the displacement of the strand complementary to the donor (forming a structure called a "D loop") and the establishment of a RAD51-bound heteroduplex species that primes DNA repair synthesis (Fig. 1). The postsynaptic stages of HR are poorly understood, but a number of proteins are involved at this stage specifcially. For example, the MCM8 and MCM9 proteins, which are related to the subunits of the hexameric MCM2-7 replicative helicase, have been implicated in postsynaptic DNA synthesis. In contrast to MCM2-7, the MCM8-9 complex is dispensable for bulk DNA replication (Griffin and Trakselis 2019). The residual DNA synthesis that is maintained in MCM2-depleted cells requires MCM8-9, but this reflects DNA repair synthesis occurring during HR-mediated repair of the high level of DSBs that occur upon MCM2 depletion (Natsume et al. 2017). Importantly, MCM8-9 was shown to act downstream from RAD51 in the HR-mediated repair of DSBs induced by nuclease overexpression, in meiotic HR, and during the HR stage of DNA interstrand cross-link (ICL) repair (Lutzmann et al. 2012;Nishimura et al. 2012;Natsume et al. 2017). The dominant mode of ICL repair is initiated by collision of replisomes with ICLs, resulting in programmed formation of DSBs that are repaired by HR. Crucially, point mutations in MCM8-9 predicted to abolish helicase activity suppress ICL repair, suggesting that helicase activity is important for HR (Nishimura et al. 2012). One explanation is that MCM8-9 might facilitate postsynaptic DNA repair synthesis by unwinding D loops and enabling extension of the invading RAD51-coated DNA end (Fig. 1).In this issue of Genes & Development, Hustedt et al. (2019) identify a new factor-HROB (HR OB-fold)-that functions closely with MCM8-9 in HR. Rationalizing that HR defects should cause sensitivity to inhibitors of ATR kinase and PARP, the investigators mined published CRISPR screens to identify new genes whose deletion sensitizes to both...

Histone chaperones control nucleosome density and chromatin structure. In yeast, the H3-H4 chaperone Spt2 controls histone deposition at active genes but its roles in metazoan chromatin structure and organismal physiology are not known. Here we identify the Caenorhabditis elegans orthologue of SPT2 (CeSPT-2) and show that its ability to bind histones H3-H4 is important for germline development and transgenerational epigenetic gene silencing, and thatspt-2mutants display signatures of a global stress response. Genome-wide profiling showed that CeSPT-2 binds to a range of highly expressed genes, and we find thatspt-2mutants have increased chromatin accessibility at these loci. We also show that human SPT2 controls the incorporation of new H3.3 into chromatin. Our work reveals roles for SPT2 in controlling chromatin structure and function in Metazoa.