To investigate the roles of substrate adhesion in cytokinesis, we established cell lines lacking paxillin （PAXB） or vineulin （VINA）, and those expressing the respective GFP fusion proteins in Dictyostelium discoid...To investigate the roles of substrate adhesion in cytokinesis, we established cell lines lacking paxillin （PAXB） or vineulin （VINA）, and those expressing the respective GFP fusion proteins in Dictyostelium discoideum. As in mammalian cells, GFP-PAXB and GFP-VINA formed focal adhesion-like complexes on the cell bottom, paxB^- cells in suspension grew normally, but on substrates, often failed to divide after regression of the furrow. The efficient cytokinesis of paxB^- cells in suspension is not because of shear forces to assist abscission, as they divided normally in static suspension culture as well. Double knockout strains lacking mhcA, which codes for myosin II, and paxB or vinA displayed more severe cytokinetic defects than each single knockout strain. In mitotic wild-type cells, GFP-PAXB was diffusely distributed on the basal membrane, hut was strikingly condensed along the polar edges in mitotic mhcA^- cells. These results are consistent with our idea that Dictyostelium displays two forms of cytokinesis, one that is contractile ringdependent and adhesion-independent, and the other that is contractile ring-independent and adhesion-dependent, and that the latter requires PAXB and VINA. Furthermore, thatpaxB- cells fail to divide normally in the presence of substrate adhesion suggests that this adhesion molecule may play additional signaling roles.展开更多
A transient postanaphase repositioning of the centriole is found to control the completion of cytokinesis. Using agreen fluorescent protein-calmodulin fusion protein as a living cell probe, we have previously found th...A transient postanaphase repositioning of the centriole is found to control the completion of cytokinesis. Using agreen fluorescent protein-calmodulin fusion protein as a living cell probe, we have previously found that calmodulin isassociated with the initiation and progression of cytokinesis. In this study, we further studied the effect of calmodulinon the repositioning of the centriole and subsequent cell cycle progression. When activity of calmodulin is inhibited, theregression of the centriole from the intercellular bridge to the cell center is blocked, and thus the completion of celldivision is repressed and two daughter cells are linked by longer cell bridge in perturbed cells. W7 treatment duringcytokinesis also results in unfinished cytokinesis and stopped G1 phase. These results suggest that calmodulin activity isrequired for centriole repositioning and can affect the completion of cytokinesis and cell cycle progression.展开更多
To probe the contributions of polar cortical cytoskeleton and the surface tension of daughter cells to intercellular bridge thinning dynamics during cytokinesis,we applied cytochalasin D（CD） or colchicine（COLC） in...To probe the contributions of polar cortical cytoskeleton and the surface tension of daughter cells to intercellular bridge thinning dynamics during cytokinesis,we applied cytochalasin D（CD） or colchicine（COLC） in a highly localized manner to polar regions of dividing normal rat kidney（NRK） cells.We observed cellular morphological changes and analyzed the intercellular bridge thinning trajectories of dividing cells with different polar cortical characteristics.Global blebbistatin（BS） application was used to obtain cells losing active contractile force groups.Our results show that locally released CD or colchicine at the polar region caused inhibition of cytokinesis before ingression.Similar treatment at phases after ingression allowed completion of cytokinesis but dramatically influenced the trajectories of intercellular bridge thinning.Disturbing single polar cortical actin induced transformation of the intercellular bridge thinning process,and polar cortical tension controlled deformation time of intercellular bridges.Our study provides a feasible framework to induce and analyze the effects of local changes in mechanical properties of cellular components on single cellular cytokinesis.展开更多
BACKGROUND Dedicator of cytokinesis 8(DOCK 8) deficiency,also known as autosomal recessive hyper immunoglobulin E(IgE) syndrome,is a combined immunodeficiency disease that was first recognized in 2009.It is caused by ...BACKGROUND Dedicator of cytokinesis 8(DOCK 8) deficiency,also known as autosomal recessive hyper immunoglobulin E(IgE) syndrome,is a combined immunodeficiency disease that was first recognized in 2009.It is caused by genetic alterations(mutations or deletions) in the DOCK 8 gene and is characterized by multiple allergies,elevated IgE levels,and susceptibility to viral and bacterial infections.Early diagnosis is critical to optimize the success of stem cell transplantation.CASE SUMMARY This study reports the case of a pediatric patient with DOCK 8 deficiency who had negative genetic testing using multiplex primary immunodeficiency(PID) panel and whole-exome sequencing(WES) with a next-generation sequencing method.He presented with chronic diarrhea and was managed as celiac disease based on previous negative workup for immunodeficiency and duodenal biopsy.He developed a generalized vesicular rash which was thought to be dermatitis herpetiformis associated with celiac disease.However,it turned out to be Eczema herpeticum based on positive herpes simplex virus from blood and lesions.The diagnosis was re-evaluated after the child was found to have multiple viral,bacterial,and parasitic co-infections(herpes simplex virus,cytomegalovirus,Epstein-Barr virus,Salmonella,and cryptosporidiosis).Re-evaluation with target gene testing with copy number variation(CNV) analysis and Multiplex Ligation Probe Amplification(MLPA) showed a large homozygous deletion in the DOCK 8 gene,confirming the diagnosis of DOCK 8 deficiency.CONCLUSION Targeted gene testing with CNV analysis might detect deletions that can be missed by WES for diagnosing patients with PID.展开更多
Cytokinesis is required for faithful division of cytoplasmic components and duplicated nuclei into two daughter cells.Midbody,a protein-dense organelle that forms at the intercellular bridge,is indispensable for succe...Cytokinesis is required for faithful division of cytoplasmic components and duplicated nuclei into two daughter cells.Midbody,a protein-dense organelle that forms at the intercellular bridge,is indispensable for success-ful cytokinesis.However,the regulatory mechanism of cytokinesis at the midbody still remains elusive.Here,we unveil a critical role for NudC-like protein 2(NudCL2),a co-chaperone of heat shock protein 90(Hsp90),in cytokinesis regulation by stabilizing regulator of chromosome condensation 2(RCC2)at the midbody in mam-malian cells.NudCL2 localizes at the midbody,and its downregulation results in cytokinesis failure,multinu-cleation,and midbody disorganization.Using iTRAQ-based quantitative proteomic analysis,we find that RCC2 levels are decreased in NudCL2 knockout(KO)cells.Moreover,Hsp90 forms a complex with NudCL2 to stabilize RCC2,which is essential for cytokinesis.RCC2 depletion mirrors phenotypes observed in NudCL2-downregulated cells.Importantly,ectopic expression of RCC2 rescues the cytokinesis defects induced by NudCL2 deletion,but not vice versa.Together,our data reveal the significance of the NudCL2/Hsp90/RCC2 pathway in cytokinesis at the midbody.展开更多
文摘To investigate the roles of substrate adhesion in cytokinesis, we established cell lines lacking paxillin （PAXB） or vineulin （VINA）, and those expressing the respective GFP fusion proteins in Dictyostelium discoideum. As in mammalian cells, GFP-PAXB and GFP-VINA formed focal adhesion-like complexes on the cell bottom, paxB^- cells in suspension grew normally, but on substrates, often failed to divide after regression of the furrow. The efficient cytokinesis of paxB^- cells in suspension is not because of shear forces to assist abscission, as they divided normally in static suspension culture as well. Double knockout strains lacking mhcA, which codes for myosin II, and paxB or vinA displayed more severe cytokinetic defects than each single knockout strain. In mitotic wild-type cells, GFP-PAXB was diffusely distributed on the basal membrane, hut was strikingly condensed along the polar edges in mitotic mhcA^- cells. These results are consistent with our idea that Dictyostelium displays two forms of cytokinesis, one that is contractile ringdependent and adhesion-independent, and the other that is contractile ring-independent and adhesion-dependent, and that the latter requires PAXB and VINA. Furthermore, thatpaxB- cells fail to divide normally in the presence of substrate adhesion suggests that this adhesion molecule may play additional signaling roles.
基金This work was supported by National Natural Science Foundation of China(Grant No.30170453)Founda tion of Ministry of Education of China for Distinguished Young Teachers(Grant No.[2001]39)awarded to Chao-Jun LI.
文摘A transient postanaphase repositioning of the centriole is found to control the completion of cytokinesis. Using agreen fluorescent protein-calmodulin fusion protein as a living cell probe, we have previously found that calmodulin isassociated with the initiation and progression of cytokinesis. In this study, we further studied the effect of calmodulinon the repositioning of the centriole and subsequent cell cycle progression. When activity of calmodulin is inhibited, theregression of the centriole from the intercellular bridge to the cell center is blocked, and thus the completion of celldivision is repressed and two daughter cells are linked by longer cell bridge in perturbed cells. W7 treatment duringcytokinesis also results in unfinished cytokinesis and stopped G1 phase. These results suggest that calmodulin activity isrequired for centriole repositioning and can affect the completion of cytokinesis and cell cycle progression.
基金supported by the National Natural Science Foundation of China (10672114)the Natural Science Foundation of Shanxi Province (2007011011)
文摘To probe the contributions of polar cortical cytoskeleton and the surface tension of daughter cells to intercellular bridge thinning dynamics during cytokinesis,we applied cytochalasin D（CD） or colchicine（COLC） in a highly localized manner to polar regions of dividing normal rat kidney（NRK） cells.We observed cellular morphological changes and analyzed the intercellular bridge thinning trajectories of dividing cells with different polar cortical characteristics.Global blebbistatin（BS） application was used to obtain cells losing active contractile force groups.Our results show that locally released CD or colchicine at the polar region caused inhibition of cytokinesis before ingression.Similar treatment at phases after ingression allowed completion of cytokinesis but dramatically influenced the trajectories of intercellular bridge thinning.Disturbing single polar cortical actin induced transformation of the intercellular bridge thinning process,and polar cortical tension controlled deformation time of intercellular bridges.Our study provides a feasible framework to induce and analyze the effects of local changes in mechanical properties of cellular components on single cellular cytokinesis.
文摘BACKGROUND Dedicator of cytokinesis 8(DOCK 8) deficiency,also known as autosomal recessive hyper immunoglobulin E(IgE) syndrome,is a combined immunodeficiency disease that was first recognized in 2009.It is caused by genetic alterations(mutations or deletions) in the DOCK 8 gene and is characterized by multiple allergies,elevated IgE levels,and susceptibility to viral and bacterial infections.Early diagnosis is critical to optimize the success of stem cell transplantation.CASE SUMMARY This study reports the case of a pediatric patient with DOCK 8 deficiency who had negative genetic testing using multiplex primary immunodeficiency(PID) panel and whole-exome sequencing(WES) with a next-generation sequencing method.He presented with chronic diarrhea and was managed as celiac disease based on previous negative workup for immunodeficiency and duodenal biopsy.He developed a generalized vesicular rash which was thought to be dermatitis herpetiformis associated with celiac disease.However,it turned out to be Eczema herpeticum based on positive herpes simplex virus from blood and lesions.The diagnosis was re-evaluated after the child was found to have multiple viral,bacterial,and parasitic co-infections(herpes simplex virus,cytomegalovirus,Epstein-Barr virus,Salmonella,and cryptosporidiosis).Re-evaluation with target gene testing with copy number variation(CNV) analysis and Multiplex Ligation Probe Amplification(MLPA) showed a large homozygous deletion in the DOCK 8 gene,confirming the diagnosis of DOCK 8 deficiency.CONCLUSION Targeted gene testing with CNV analysis might detect deletions that can be missed by WES for diagnosing patients with PID.
基金supported by the National Natural Science Foundation of China(Nos.32070709,32270771,and U21A20197)the National Key Research and Development Program of China(Nos.2019YFA0802202)the Higher Education Discipline Innovation Project(also known as the111 Project)(Nos.B13026).
文摘Cytokinesis is required for faithful division of cytoplasmic components and duplicated nuclei into two daughter cells.Midbody,a protein-dense organelle that forms at the intercellular bridge,is indispensable for success-ful cytokinesis.However,the regulatory mechanism of cytokinesis at the midbody still remains elusive.Here,we unveil a critical role for NudC-like protein 2(NudCL2),a co-chaperone of heat shock protein 90(Hsp90),in cytokinesis regulation by stabilizing regulator of chromosome condensation 2(RCC2)at the midbody in mam-malian cells.NudCL2 localizes at the midbody,and its downregulation results in cytokinesis failure,multinu-cleation,and midbody disorganization.Using iTRAQ-based quantitative proteomic analysis,we find that RCC2 levels are decreased in NudCL2 knockout(KO)cells.Moreover,Hsp90 forms a complex with NudCL2 to stabilize RCC2,which is essential for cytokinesis.RCC2 depletion mirrors phenotypes observed in NudCL2-downregulated cells.Importantly,ectopic expression of RCC2 rescues the cytokinesis defects induced by NudCL2 deletion,but not vice versa.Together,our data reveal the significance of the NudCL2/Hsp90/RCC2 pathway in cytokinesis at the midbody.
