Dysregulation of cytoskeletal proteins has been found in response to DNA damage stress,yet the functional role of cytoskeletal proteins in DNA repair remained unexplored.Here,we found that DNA-damaging agents induced ...Dysregulation of cytoskeletal proteins has been found in response to DNA damage stress,yet the functional role of cytoskeletal proteins in DNA repair remained unexplored.Here,we found that DNA-damaging agents induced substantial upregulation of smooth muscle-specific cytoskeletal protein smoothelin(SMTN)in colorectal cancer(CRC)cells.Silencing SMTN abrogated G2/M arrest,exacerbated DNA damage,and markedly enhanced the chemosensitivity of CRC cells to various DNA-damaging agents.Notably,SMTN could rapidly accumulate at DNA damage sites within 1 min after laser irradiation,which was indispensable for the initiation of homologous recombination(HR)repair.Mechanistically,SMTN stabilized RAD51 by disrupting its interaction with its E3 ubiquitin ligase HUWE1,thereby maintaining the process of HR repair.To explore the therapeutic role of SMTN,customized cell membrane infused biomimetic liposomes were constructed to ensure rapid delivery of SMTN siRNA specifically into HCT-116 cells,yielding significantly enhanced anti-cancer effects of irinotecan and fuzuloparib both in vitro and in vivo.To summarize,our findings revealed a novel function of SMTN in DNA damage repair and provided a therapeutic strategy of targeting SMTN to enhance the efficacy of DNA damage agents.展开更多
Single-base editors,including cytosine base editors(CBEs)and adenine base editors(ABEs),facilitate accurate C·G to T·A and A·T to G·C,respectively,holding promise for the precise modeling and treat...Single-base editors,including cytosine base editors(CBEs)and adenine base editors(ABEs),facilitate accurate C·G to T·A and A·T to G·C,respectively,holding promise for the precise modeling and treatment of human hereditary disorders.Efficient base editing and expanded base conversion range have been achieved in human cells through base editors fusing with Rad51 DNA binding domain(Rad51DBD),such as hyA3A-BE4max.Here,we show that hyA3A-BE4max catalyzes C-to-T substitution in the zebrafish genome and extends editing positions(C_(12)-C_(16))proximal to the protospacer adjacent motif.We develop a codon-optimized counterpart zhyA3A-CBE5,which exhibits substantially high C-to-T conversion with 1.59-to 3.50-fold improvement compared with the original hyA3A-BE4max.With these tools,disease-relevant hereditary mutations can be more efficaciously generated in zebrafish.We introduce human genetic mutation rpl11^(Q42*)and abcc6a^(R1463C) by zhyA3A-CBE5 in zebrafish,mirroring Diamond-Blackfan anemia and Pseudoxanthoma Elasticum,respectively.Our study expands the base editing platform targeting the zebrafish genomic landscape and the application of single-base editors for disease modeling and gene function study.展开更多
REV1是跨损伤聚合酶Y家族的重要成员之一,它不仅作为支架蛋白介导Y家族聚合酶招募至损伤位点完成跨损伤DNA合成(translesion DNA synthesis, TLS),还可利用自身的dCMP转移酶活性在一些损伤位点对侧整合dCMP参与TLS。此外,REV1也被报导...REV1是跨损伤聚合酶Y家族的重要成员之一,它不仅作为支架蛋白介导Y家族聚合酶招募至损伤位点完成跨损伤DNA合成(translesion DNA synthesis, TLS),还可利用自身的dCMP转移酶活性在一些损伤位点对侧整合dCMP参与TLS。此外,REV1也被报导参与调控同源重组修复。为进一步探讨REV1互作蛋白RAD51和RAD51C在其参与的同源重组修复通路中的调控作用,本研究采用脉冲氮激光微辐射实验,发现RAD51可调控REV1到双链断裂位点的募集。同时,免疫荧光实验结果证明REV1也反过来影响RAD51应答CPT损伤。然而敲低RAD51C并不影响REV1到DNA双链断裂位点的招募。结果表明,REV1和RAD51在HR通路中存在彼此相互调控的关系。展开更多
基金supported by National Natural Science Foundation(82073297,China)Department of Science and Technology of Zhejiang Province Key R&D Projects(2022C03155 and 2023C03112,China)+2 种基金Zhejiang Provincial Program for the Cultivation of High-level Leading Health Talents(2020-18,China)Clinical Pharmacy of Hangzhou Medical Key Discipline(2021-21-16,China)Huzhou City Public Application Research Project(2019GYB63,China)。
文摘Dysregulation of cytoskeletal proteins has been found in response to DNA damage stress,yet the functional role of cytoskeletal proteins in DNA repair remained unexplored.Here,we found that DNA-damaging agents induced substantial upregulation of smooth muscle-specific cytoskeletal protein smoothelin(SMTN)in colorectal cancer(CRC)cells.Silencing SMTN abrogated G2/M arrest,exacerbated DNA damage,and markedly enhanced the chemosensitivity of CRC cells to various DNA-damaging agents.Notably,SMTN could rapidly accumulate at DNA damage sites within 1 min after laser irradiation,which was indispensable for the initiation of homologous recombination(HR)repair.Mechanistically,SMTN stabilized RAD51 by disrupting its interaction with its E3 ubiquitin ligase HUWE1,thereby maintaining the process of HR repair.To explore the therapeutic role of SMTN,customized cell membrane infused biomimetic liposomes were constructed to ensure rapid delivery of SMTN siRNA specifically into HCT-116 cells,yielding significantly enhanced anti-cancer effects of irinotecan and fuzuloparib both in vitro and in vivo.To summarize,our findings revealed a novel function of SMTN in DNA damage repair and provided a therapeutic strategy of targeting SMTN to enhance the efficacy of DNA damage agents.
基金supported by grants from Ministry of Science and Technology of the People's Republic of China(2018YFA0801004 and 2018YFA0800103)the National Natural Science Foundation of China(NSFC31530044,NSFC31970780,NSFC82202056).
文摘Single-base editors,including cytosine base editors(CBEs)and adenine base editors(ABEs),facilitate accurate C·G to T·A and A·T to G·C,respectively,holding promise for the precise modeling and treatment of human hereditary disorders.Efficient base editing and expanded base conversion range have been achieved in human cells through base editors fusing with Rad51 DNA binding domain(Rad51DBD),such as hyA3A-BE4max.Here,we show that hyA3A-BE4max catalyzes C-to-T substitution in the zebrafish genome and extends editing positions(C_(12)-C_(16))proximal to the protospacer adjacent motif.We develop a codon-optimized counterpart zhyA3A-CBE5,which exhibits substantially high C-to-T conversion with 1.59-to 3.50-fold improvement compared with the original hyA3A-BE4max.With these tools,disease-relevant hereditary mutations can be more efficaciously generated in zebrafish.We introduce human genetic mutation rpl11^(Q42*)and abcc6a^(R1463C) by zhyA3A-CBE5 in zebrafish,mirroring Diamond-Blackfan anemia and Pseudoxanthoma Elasticum,respectively.Our study expands the base editing platform targeting the zebrafish genomic landscape and the application of single-base editors for disease modeling and gene function study.
文摘REV1是跨损伤聚合酶Y家族的重要成员之一,它不仅作为支架蛋白介导Y家族聚合酶招募至损伤位点完成跨损伤DNA合成(translesion DNA synthesis, TLS),还可利用自身的dCMP转移酶活性在一些损伤位点对侧整合dCMP参与TLS。此外,REV1也被报导参与调控同源重组修复。为进一步探讨REV1互作蛋白RAD51和RAD51C在其参与的同源重组修复通路中的调控作用,本研究采用脉冲氮激光微辐射实验,发现RAD51可调控REV1到双链断裂位点的募集。同时,免疫荧光实验结果证明REV1也反过来影响RAD51应答CPT损伤。然而敲低RAD51C并不影响REV1到DNA双链断裂位点的招募。结果表明,REV1和RAD51在HR通路中存在彼此相互调控的关系。
