CtBP-interacting protein(CtIP)is known for its multifaceted roles in DNA repair and genomic stability,directing the homologous recombination-mediated DNA double-stranded break repair pathway via DNA end resection,an e...CtBP-interacting protein(CtIP)is known for its multifaceted roles in DNA repair and genomic stability,directing the homologous recombination-mediated DNA double-stranded break repair pathway via DNA end resection,an essential error-free repair process vital for genome stability.Mammalian oocytes are highly prone to DNA damage accumulation due to prolonged G2/prophase arrest.Here,we explore the functions of CtIP in meiotic cell cycle regulation via a mouse oocyte model.Depletion of CtIP by siRNA injection results in delayed germinal vesicle breakdown and failed polar body extrusion.Mechanistically,CtIP deficiency increases DNA damage and decreases the expression and nuclear entry of CCNB1,resulting in marked impairment of meiotic resumption,which can be rescued by exogenous CCNB1 overexpression.Furthermore,depletion of CtIP disrupts microtubule-organizing centers coalescence at spindle poles as indicated by failed accumulation ofγ-tubulin,p-Aurora kinase A,Kif2A,and TPX2,leading to abnormal spindle assembly and prometaphase arrest.These results provide valuable insights into the important roles of CtIP in the G2/M checkpoint and spindle assembly in mouse oocyte meiotic cell cycle regulation.展开更多
Background:CK2(casein kinase 2)is a serine/threonine-selective protein kinase that has been involved in a variety of cellular processes such as DNA repair,cell cycle control and circadian rhythm regulation.However,its...Background:CK2(casein kinase 2)is a serine/threonine-selective protein kinase that has been involved in a variety of cellular processes such as DNA repair,cell cycle control and circadian rhythm regulation.However,its functional roles in oocyte meiosis have not been fully determined.Results:We report that CK2 is essential for porcine oocyte meiotic maturation by regulating spindle assembly checkpoint(SAC).Immunostaining and immunoblotting analysis showed that CK2 was constantly expressed and located on the chromosomes during the entire oocyte meiotic maturation.Inhibition of CK2 activity by its selective inhibitor CX-4945 impaired the first polar body extrusion and arrested oocytes at M I stage,accompanied by the presence of BubR1 at kinetochores,indicative of activated SAC.In addition,we found that spindle/chromosome structure was disrupted in CK2-inhibited oocytes due to the weakened microtubule stability,which is a major cause resulting in the activation of SAC.Last,we found that the level DNA damage as assessed byγH2A.X staining was considerably elevated when CK2 was inhibited,suggesting that DNA damage might be another critical factor leading to the SAC activation and meiotic failure of oocytes.Conclusions:Our findings demonstrate that CK2 promotes the porcine oocyte maturation by ensuring normal spindle assembly and DNA damage repair.展开更多
Error-free mitosis depends on accurate chromosome attachment to spindle microtubules,which is monitored by the spindle assembly checkpoint(SAC)signaling.As an upstream factor of SAC,the precise and dynamic kinetochore...Error-free mitosis depends on accurate chromosome attachment to spindle microtubules,which is monitored by the spindle assembly checkpoint(SAC)signaling.As an upstream factor of SAC,the precise and dynamic kinetochore localization of Mps1 kinase is critical for initiating and silencing SAC signaling.However,the underlying molecular mechanism remains elusive.Here,we demonstrated that the multisite interactions between Mps1 and Ndc80 complex(Ndc80C)govern Mps1 kinetochore targeting.Importantly,we identified direct interaction between Mps1 tetratricopeptide repeat domain and Ndc80C.We further identified that Mps1 C-terminal fragment,which contains the protein kinase domain and C-tail,enhances Mps1 kinetochore localization.Mechanistically,Mps1 C-terminal fragment mediates its dimerization.Perturbation of C-tail attenuates the kinetochore targeting and activity of Mps1,leading to aberrant mitosis due to compromised SAC function.Taken together,our study highlights the importance of Mps1 dimerization and multisite interactions with Ndc80C in enabling responsive SAC signaling.展开更多
Loss or gain of chromosomes,a condition known as aneuploidy,is a common feature of tumor cells and has therefore been proposed as the driving force for tumorigenesis.Such chromosomal instability can arise during mitos...Loss or gain of chromosomes,a condition known as aneuploidy,is a common feature of tumor cells and has therefore been proposed as the driving force for tumorigenesis.Such chromosomal instability can arise during mitosis as a result of mis-segregation of the duplicated sister chromatids to the two daughter cells.In normal cells,missegregation is usually prevented by the spindle assembly checkpoint(SAC),a sophisticated surveillance mechanism that inhibits mitotic exit until all chromosomes have successfully achieved bipolar attachment to spindle microtubules.Complete abrogation of SAC activity is lethal to normal as well as to tumor cells,as a consequence of massive chromosome mis-segregation.Importantly,many human aneuploid tumor cells exhibit a weakened SAC activity that allows them to tolerate gains or losses of a small number of chromosomes;and interfering with this SAC residual activity may constitute a suitable strategy to kill cancer cells.This review focuses on the potential link between SAC and tumorigenesis,and the therapeutic strategy to target the SAC for cancer treatment.展开更多
The microtubule organizing centers(MTOCs)of human and mouse oocytes are essential for meiotic spindle assembly and for ensuring precise chromosome segregations.Previous studies mainly focus on investigating MTOCs chan...The microtubule organizing centers(MTOCs)of human and mouse oocytes are essential for meiotic spindle assembly and for ensuring precise chromosome segregations.Previous studies mainly focus on investigating MTOCs changes in metaphase I oocyte.However,the detailed dynamic changes and underlying mechanisms of the MTOCs in germinal vesicle(GV)oocytes—a stage that early events of MTOC maturation happened—remain unclear.Here we explored the dynamics of MTOCs maturation in human and mouse GV oocytes and found that MTOCs maturation is a largely conserved process,consisting of two tightly coupled processes referred to as MTOCs activation and migration.We found that cytoskeleton associated protein 5(CKAP5)and transforming acidic coiled-coil containing protein 3(TACC3)play key roles in MTOCs maturation in oocytes.The activation of the MTOCs is a prerequisite for migration initiation,and the migration of the MTOCs is facilitated by dynein/dynactin in oocytes.The disruption of MTOC maturation resulted in spindle assembly failure.Importantly,impaired MTOCs maturation is associated with the physiological aging of oocytes.This study provides a comprehensive understanding of MTOCs dynamics in human and mouse oocytes.展开更多
The Bub1 and BubR1 kinetochore proteins support proper chromosome segregation and mitotic checkpoint activity. Bub1 and BubR1 are paralogs with Bub1 being a kinase, while BubR1 localizes the PP2A-B56 protein phosphata...The Bub1 and BubR1 kinetochore proteins support proper chromosome segregation and mitotic checkpoint activity. Bub1 and BubR1 are paralogs with Bub1 being a kinase, while BubR1 localizes the PP2A-B56 protein phosphatase to kinetochores in humans. Whether this spatial separation of kinase and phosphatase activity is important is unclear as some organisms integrate both activities into one Bub protein. Here, we engineer human Bub1 and BubR1 proteins integrating kinase and phosphatase activities into one protein and show that these do not support normal mitotic progression. A Bub1–PP2A-B56 complex can support chromosome alignment but results in impairment of the checkpoint due to dephosphorylation of the Mad1 binding site in Bub1. Furthermore, a chimeric BubR1 protein containing the Bub1 kinase domain induces delocalized H2ApT120 phosphorylation, resulting in the reduction of centromeric hSgo2 and chromosome segregation errors. Collectively, these results argue that the spatial separation of kinase and phosphatase activities within the Bub complex is required for balancing its functions in the checkpoint and chromosome alignment.展开更多
Chromosome segregation is a tightly regulated process through which duplicated genetic materials are equally partitioned into daughter cells. During the past decades, tremendous efforts have been made to understand th...Chromosome segregation is a tightly regulated process through which duplicated genetic materials are equally partitioned into daughter cells. During the past decades, tremendous efforts have been made to understand the molecular mechanism of chromosome segregation using animals and yeasts as model systems. Recently, new insights into chromosome segregation have gradually emerged using trypanosome, an early branching parasitic protozoan, as a model organism. To uncover the unique aspects of chromosome segregation in trypanosome, which potentially could serve as new drug targets for anti-trypanosome chemotherapy, it is necessary to perform a comparative analysis of the chromosome segregation machinery between trypanosome and its human host. Here, we briefly review the current knowledge about chromosome segregation in human and Trypanosoma brucei, with a focus on the regulation of cohesin and securin degradation triggered by the activation of the anaphase promoting complex/cyclosome （APC/C）. We also include yeasts in our comparative analysis since some of the original discoveries were made using budding and fission yeasts as the model organisms and, therefore, these could provide hints about the evolution of the machinery. We highlight both common and unique features in these model systems and also provide perspectives for future research in trypanosome.展开更多
基金supported by National Natural Science Foundation of China(32570854)Science and Technology Program of Guangzhou,China(2023A03J0258)Guangdong Basic and Applied Basic Research Foundation,China(2023B1515120027)。
文摘CtBP-interacting protein(CtIP)is known for its multifaceted roles in DNA repair and genomic stability,directing the homologous recombination-mediated DNA double-stranded break repair pathway via DNA end resection,an essential error-free repair process vital for genome stability.Mammalian oocytes are highly prone to DNA damage accumulation due to prolonged G2/prophase arrest.Here,we explore the functions of CtIP in meiotic cell cycle regulation via a mouse oocyte model.Depletion of CtIP by siRNA injection results in delayed germinal vesicle breakdown and failed polar body extrusion.Mechanistically,CtIP deficiency increases DNA damage and decreases the expression and nuclear entry of CCNB1,resulting in marked impairment of meiotic resumption,which can be rescued by exogenous CCNB1 overexpression.Furthermore,depletion of CtIP disrupts microtubule-organizing centers coalescence at spindle poles as indicated by failed accumulation ofγ-tubulin,p-Aurora kinase A,Kif2A,and TPX2,leading to abnormal spindle assembly and prometaphase arrest.These results provide valuable insights into the important roles of CtIP in the G2/M checkpoint and spindle assembly in mouse oocyte meiotic cell cycle regulation.
基金This work was supported by the National Natural Science Foundation of China(31822053,31900592)Natural Science Foundation of Jiangsu Province(SBK2019043265)China Postdoctoral Science Foundation(2019M651849).
文摘Background:CK2(casein kinase 2)is a serine/threonine-selective protein kinase that has been involved in a variety of cellular processes such as DNA repair,cell cycle control and circadian rhythm regulation.However,its functional roles in oocyte meiosis have not been fully determined.Results:We report that CK2 is essential for porcine oocyte meiotic maturation by regulating spindle assembly checkpoint(SAC).Immunostaining and immunoblotting analysis showed that CK2 was constantly expressed and located on the chromosomes during the entire oocyte meiotic maturation.Inhibition of CK2 activity by its selective inhibitor CX-4945 impaired the first polar body extrusion and arrested oocytes at M I stage,accompanied by the presence of BubR1 at kinetochores,indicative of activated SAC.In addition,we found that spindle/chromosome structure was disrupted in CK2-inhibited oocytes due to the weakened microtubule stability,which is a major cause resulting in the activation of SAC.Last,we found that the level DNA damage as assessed byγH2A.X staining was considerably elevated when CK2 was inhibited,suggesting that DNA damage might be another critical factor leading to the SAC activation and meiotic failure of oocytes.Conclusions:Our findings demonstrate that CK2 promotes the porcine oocyte maturation by ensuring normal spindle assembly and DNA damage repair.
基金supported by the National Key R&D Program of China(2017YFA 0102900 and 2017 YFA 0503600)the National Natural Science Fondation of China(31671407 and 31871359 to Z.D.+4 种基金31621002,31430054,91854203,and 31320103904 to X.Y.31301099 and 21672201 to X.G.31471275 to D.W.),Strategic Priority Research Program of the Chinese Academy of Sciences(XDB19040000)Chinese Academy of Sciences Center for Excellence in Molecular Cell Science(2015 HSC-UE010)MOE Innovative Team(IRT_17R102).
文摘Error-free mitosis depends on accurate chromosome attachment to spindle microtubules,which is monitored by the spindle assembly checkpoint(SAC)signaling.As an upstream factor of SAC,the precise and dynamic kinetochore localization of Mps1 kinase is critical for initiating and silencing SAC signaling.However,the underlying molecular mechanism remains elusive.Here,we demonstrated that the multisite interactions between Mps1 and Ndc80 complex(Ndc80C)govern Mps1 kinetochore targeting.Importantly,we identified direct interaction between Mps1 tetratricopeptide repeat domain and Ndc80C.We further identified that Mps1 C-terminal fragment,which contains the protein kinase domain and C-tail,enhances Mps1 kinetochore localization.Mechanistically,Mps1 C-terminal fragment mediates its dimerization.Perturbation of C-tail attenuates the kinetochore targeting and activity of Mps1,leading to aberrant mitosis due to compromised SAC function.Taken together,our study highlights the importance of Mps1 dimerization and multisite interactions with Ndc80C in enabling responsive SAC signaling.
基金H.B.was supported by grant#04-GBMC-CICS-09,from CESPU–Cooperativa de Ensino Superior Politécnico e Universitário.
文摘Loss or gain of chromosomes,a condition known as aneuploidy,is a common feature of tumor cells and has therefore been proposed as the driving force for tumorigenesis.Such chromosomal instability can arise during mitosis as a result of mis-segregation of the duplicated sister chromatids to the two daughter cells.In normal cells,missegregation is usually prevented by the spindle assembly checkpoint(SAC),a sophisticated surveillance mechanism that inhibits mitotic exit until all chromosomes have successfully achieved bipolar attachment to spindle microtubules.Complete abrogation of SAC activity is lethal to normal as well as to tumor cells,as a consequence of massive chromosome mis-segregation.Importantly,many human aneuploid tumor cells exhibit a weakened SAC activity that allows them to tolerate gains or losses of a small number of chromosomes;and interfering with this SAC residual activity may constitute a suitable strategy to kill cancer cells.This review focuses on the potential link between SAC and tumorigenesis,and the therapeutic strategy to target the SAC for cancer treatment.
基金supported by the National Natural Science Foundation of China(82325021,82288102,32130029,82422033,82271685,82171643,and 82171685)the Shanghai Rising-Star Program(23QA1401800)+3 种基金the New Cornerstone Science Foundation through the XPLORER PRIZE,the Fund of Fudan University and Cao’ejiang Basic Research(24FCB01)the SANS Exploration Scholarthe Fund for Excellent Young Scholars of Shanghai Ninth People’s Hospital,Shanghai Jiao Tong University School of Medicine(JYYQ004)the Shanghai JIAI Genetics and IVF Institute Found(JA-2025033).
文摘The microtubule organizing centers(MTOCs)of human and mouse oocytes are essential for meiotic spindle assembly and for ensuring precise chromosome segregations.Previous studies mainly focus on investigating MTOCs changes in metaphase I oocyte.However,the detailed dynamic changes and underlying mechanisms of the MTOCs in germinal vesicle(GV)oocytes—a stage that early events of MTOC maturation happened—remain unclear.Here we explored the dynamics of MTOCs maturation in human and mouse GV oocytes and found that MTOCs maturation is a largely conserved process,consisting of two tightly coupled processes referred to as MTOCs activation and migration.We found that cytoskeleton associated protein 5(CKAP5)and transforming acidic coiled-coil containing protein 3(TACC3)play key roles in MTOCs maturation in oocytes.The activation of the MTOCs is a prerequisite for migration initiation,and the migration of the MTOCs is facilitated by dynein/dynactin in oocytes.The disruption of MTOC maturation resulted in spindle assembly failure.Importantly,impaired MTOCs maturation is associated with the physiological aging of oocytes.This study provides a comprehensive understanding of MTOCs dynamics in human and mouse oocytes.
基金supported by the National Natural Science Foundation of China(31970666)Taishan Scholar Project(tsqn201812054)from Shandong,China+3 种基金Work at the Novo Nordisk Foundation Center for Protein Research was supported by the grant NNF14CC0001J.N.is supported by grants from the Danish Cancer Society(R269-A15586-B17)Independent Research Fund Denmark(8021-00101B and 0134-00199B)Novo Nordisk Foundation(NNF20OC0065098).
文摘The Bub1 and BubR1 kinetochore proteins support proper chromosome segregation and mitotic checkpoint activity. Bub1 and BubR1 are paralogs with Bub1 being a kinase, while BubR1 localizes the PP2A-B56 protein phosphatase to kinetochores in humans. Whether this spatial separation of kinase and phosphatase activity is important is unclear as some organisms integrate both activities into one Bub protein. Here, we engineer human Bub1 and BubR1 proteins integrating kinase and phosphatase activities into one protein and show that these do not support normal mitotic progression. A Bub1–PP2A-B56 complex can support chromosome alignment but results in impairment of the checkpoint due to dephosphorylation of the Mad1 binding site in Bub1. Furthermore, a chimeric BubR1 protein containing the Bub1 kinase domain induces delocalized H2ApT120 phosphorylation, resulting in the reduction of centromeric hSgo2 and chromosome segregation errors. Collectively, these results argue that the spatial separation of kinase and phosphatase activities within the Bub complex is required for balancing its functions in the checkpoint and chromosome alignment.
文摘Chromosome segregation is a tightly regulated process through which duplicated genetic materials are equally partitioned into daughter cells. During the past decades, tremendous efforts have been made to understand the molecular mechanism of chromosome segregation using animals and yeasts as model systems. Recently, new insights into chromosome segregation have gradually emerged using trypanosome, an early branching parasitic protozoan, as a model organism. To uncover the unique aspects of chromosome segregation in trypanosome, which potentially could serve as new drug targets for anti-trypanosome chemotherapy, it is necessary to perform a comparative analysis of the chromosome segregation machinery between trypanosome and its human host. Here, we briefly review the current knowledge about chromosome segregation in human and Trypanosoma brucei, with a focus on the regulation of cohesin and securin degradation triggered by the activation of the anaphase promoting complex/cyclosome （APC/C）. We also include yeasts in our comparative analysis since some of the original discoveries were made using budding and fission yeasts as the model organisms and, therefore, these could provide hints about the evolution of the machinery. We highlight both common and unique features in these model systems and also provide perspectives for future research in trypanosome.
