Dpb4 promotes resection of DNA double-strand breaks and checkpoint activation by acting in two different protein complexes

Erika Casari,Elisa Gobbini,Marco Gnugnoli,Maria Pia Longhese,Marco Mangiagalli,Michela Clerici

doi:10.1038/s41467-021-25090-9

Erika Casari, Elisa Gobbini + Show 4 more

Open Access

https://doi.org/10.1038/s41467-021-25090-9

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Journal: Nature Communications	Publication Date: Aug 6, 2021
Citations: 10	License type: open-access

Affiliation: University of Milano-Bicocca

Abstract

Budding yeast Dpb4 (POLE3/CHRAC17 in mammals) is a highly conserved histone fold protein that is shared by two protein complexes: the chromatin remodeler ISW2/hCHRAC and the DNA polymerase ε (Pol ε) holoenzyme. In Saccharomyces cerevisiae, Dpb4 forms histone-like dimers with Dls1 in the ISW2 complex and with Dpb3 in the Pol ε complex. Here, we show that Dpb4 plays two functions in sensing and processing DNA double-strand breaks (DSBs). Dpb4 promotes histone removal and DSB resection by interacting with Dls1 to facilitate the association of the Isw2 ATPase to DSBs. Furthermore, it promotes checkpoint activation by interacting with Dpb3 to facilitate the association of the checkpoint protein Rad9 to DSBs. Persistence of both Isw2 and Rad9 at DSBs is enhanced by the A62S mutation that is located in the Dpb4 histone fold domain and increases Dpb4 association at DSBs. Thus, Dpb4 exerts two distinct functions at DSBs depending on its interactors.

Highlights

Budding yeast Dpb[4] (POLE3/CHRAC17 in mammals) is a highly conserved histone fold protein that is shared by two protein complexes: the chromatin remodeler ISW2/hCHRAC and the DNA polymerase ε (Pol ε) holoenzyme
To understand whether the dpb4-A62S mutation exacerbates the DNA damage sensitivity of tel1Δ and sae2Δ cells by disrupting Dpb[4] function, we analyzed the effect of DPB4 deletion. dpb4Δ tel1Δ and dpb4Δ sae2Δ cells were less sensitive to CPT than dpb4-A62S tel1Δ and dpb4-A62S sae2Δ cells, respectively (Fig. 1a, b), suggesting that the synthetic effect caused by the dpb4-A62S allele is not due to loss of Dpb[4] function
The DPB4 deletion increased less severely the DNA damage sensitivity of tel1Δ and sae2Δ cells compared to dpb4A62S, dpb4Δ cells were more sensitive than dpb4-A62S cells to a high CPT dose, and to phleomycin and methyl methanesulfonate (MMS) (Fig. 1c)

Summary

Introduction

Budding yeast Dpb[4] (POLE3/CHRAC17 in mammals) is a highly conserved histone fold protein that is shared by two protein complexes: the chromatin remodeler ISW2/hCHRAC and the DNA polymerase ε (Pol ε) holoenzyme. DSB ends undergo nucleolytic degradation (resection), generating 3′-ended single-stranded DNA (ssDNA) tails that can invade the undamaged homologous DNA template[3]. DSB resection is initiated by the binding to the DSB ends of the evolutionarily conserved Mre11-Rad50-Xrs2/NBS1 (MRX/N) complex[4]. The Sae[2] protein (CtIP in mammals) activates a latent endonuclease activity of Mre[11], which cleaves the 5′-terminated strands at both DNA ends[5]. The resulting nick generates an entry site for both Mre[11], which degrades back toward the DSB end in a 3′–5′.

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