HPF1-dependent PARP activation promotes LIG3-XRCC1-mediated backup pathway of Okazaki fragment ligation

DNA ligase 1 (LIG1) is known as the major DNA ligase responsible for Okazaki fragment joining. Recent studies have implicated LIG3 complexed with XRCC1 as an alternative player in Okazaki fragment joining in cases where LIG1 is not functional, although the underlying mechanisms are largely unknown. Here, using a cell-free system derived from Xenopus egg extracts, we demonstrated the essential role of PARP1-HPF1 in LIG3-dependent Okazaki fragment joining. We found that Okazaki fragments were eventually ligated even in the absence of LIG1, employing in its place LIG3-XRCC1, which was recruited onto chromatin. Concomitantly, LIG1 deficiency induces ADP-ribosylation of histone H3 in a PARP1-HPF1-dependent manner. The depletion of PARP1 or HPF1 resulted in a failure to recruit LIG3 onto chromatin and a subsequent failure in Okazaki fragment joining in LIG1-depleted extracts. Importantly, Okazaki fragments were not ligated at all when LIG1 and XRCC1 were co-depleted. Our results suggest that a unique form of ADP-ribosylation signaling promotes the recruitment of LIG3 on chromatin and its mediation of Okazaki fragment joining as a backup system for LIG1 perturbation

Two distinct modes of DNMT1 recruitment ensure stable maintenance DNA methylation

Stable inheritance of DNA methylation is critical for maintaining differentiated phenotypes in multicellular organisms. We have recently identified dual mono-ubiquitylation of histone H3 (H3Ub2) by UHRF1 as an essential mechanism to recruit DNMT1 to chromatin. Here, we show that PCNA-associated factor 15 (PAF15) undergoes UHRF1-dependent dual mono-ubiquitylation (PAF15Ub2) on chromatin in a DNA replication-coupled manner. This event will, in turn, recruit DNMT1. During early S-phase, UHRF1 preferentially ubiquitylates PAF15, whereas H3Ub2 predominates during late S-phase. H3Ub2 is enhanced under PAF15 compromised conditions, suggesting that H3Ub2 serves as a backup for PAF15Ub2. In mouse ES cells, loss of PAF15Ub2 results in DNA hypomethylation at early replicating domains. Together, our results suggest that there are two distinct mechanisms underlying replication timing-dependent recruitment of DNMT1 through PAF15Ub2 and H3Ub2, both of which are prerequisite for high fidelity DNA methylation inheritance

The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target

Sputtering Yields of Si Bombarded with 10-540-keV C60 Ions

Sputtering yields of Si have been measured for C60 ions in the energy range from 10 to 540 keV, where the nuclear stopping is dominant, by measuring thickness change of a pre-amorphized layer with conventional Rutherford-backscattering spectroscopy. The measured sputtering yield shows the maximum, which is approximately 600 Si/C60, around 100 keV. Comparing with the sputtering yields for a monatomic ion calculated both based on the linear-collision-cascade theory of Sigmund and using the SRIM2008 code, nonlinear effect on the sputtering yield has been observed. The nonlinear effect depends on the energy of C60 ions: it is very large around the energies where the sputtering yield has the maximum and hardly observed at 10 keV

Development of a highly-intense negative C60 ion source for a tandem accelerator and its applications

　高崎量子応用研究所イオン照射研究施設(TIARA)におけるMeV級C60イオンビームを安定に利用するための、タンデム加速器用の負イオン源の開発について報告した。次にこのイオン源開発により可能になった研究として、MeV級C60イオンビーム照射を利用した、金のスパッタリング収量の測定をはじめとしたいくつかの成果を紹介した。23rd International Workshop on Inelastic Ion-Surface Collisions (IISC-23

A Comparative Study on Lethal Effect of Monomer and Cluster Proton Ion Beams in Bacterial Spores

クラスターイオンは、複数の原子がnm距離に近接した状態で物質に入射することから、材料表面の改質や分析では単原子イオンに比べて特異なエネルギー付与を示すことが知られている。しかしながら、生物試料における研究は実施されていなかった。我々は、枯草菌の胞子を生物試料のモデルとし、クラスターイオンの生物効果の特徴について明らかにすることを目的とした研究を行っている。原子あたり2MeVの炭素イオンの致死効果を評価した結果、LETが非常に高いために1原子のヒットで致死するため、致死効果に関するクラスター化の効果は確認されないことがわかった。そこで、炭素よりも低いLETを持つプロトンのクラスターイオンの致死効果を評価した。2 MeV H2は1MeV Hに比べて原子あたりの致死効果が低く、負のクラスター効果を示す可能性が示唆された。これは、2つのプロトンが近接して通過するために、同じ遺伝子に影響する確率が高いことによると解釈できる。340 keV Hは2MeV H2と粒子あたりで同じLETを持つが、致死効果は340 keV Hの方が明らかに高かった。これは、イオントラック中心付近でのエネルギー密度が致死効果に大きく影響することによると考えられる