Actin cytoskeleton plays crucial roles in various cellular functions.Extracellular matrix(ECM)can modulate cell morphology by remodeling the internal cytoskeleton.To define how geometry of ECM regulates the organizati...Actin cytoskeleton plays crucial roles in various cellular functions.Extracellular matrix(ECM)can modulate cell morphology by remodeling the internal cytoskeleton.To define how geometry of ECM regulates the organization of actin cytoskeleton,we plated individual NIH 3T3 cells on micropatterned substrates with distinct shapes and sizes.It was found that the stress fibers could form along the nonadhesive edges of T-shaped pattern,but were absent from the opening edge of V-shaped pattern,indicating that the organization of actin cytoskeleton was dependent on the mechanical environment.Furthermore,a secondary actin ring was observed on 50μm circular pattern while did not appear on 30μm and 40μm pattern,showing a size-dependent organization of actin cytoskeleton.Finally,osteoblasts,MDCK and A549 cells exhibited distinct organization of actin cytoskeleton on T-shaped pattern,suggesting a cell-type specificity in arrangement of actin cytoskeleton.Together,our findings brought novel insight into the organization of actin cytoskeleton on micropatterned environments.展开更多
As the indispensable oxygen-transporting cells,erythrocytes exhibit extreme deformability and amazing stability as they are subject to huge reversible shear stress and extrusion force during massive circulation in the...As the indispensable oxygen-transporting cells,erythrocytes exhibit extreme deformability and amazing stability as they are subject to huge reversible shear stress and extrusion force during massive circulation in the body.The unique architecture of spectrin-actin-based membraneskeleton is considered to be responsible for such excellent mechanical properties of erythrocytes.Although erythrocytes have been recognized for more than 300 years,myriad questions about membrane-skeleton constantly attract people's attention.Here,we summarize the kinds of distinctive single-cell and single-molecule techniques that were used to investigate the structure and function of erythrocyte membrane-skeleton at macro and micro levels.展开更多
Cells are highly sensitive to their geometrical and mechanical microenvironment that directly regulate cell shape,cytoskeleton and organelle,as well as the nucleus morphology and genetic expression.The emerging two-di...Cells are highly sensitive to their geometrical and mechanical microenvironment that directly regulate cell shape,cytoskeleton and organelle,as well as the nucleus morphology and genetic expression.The emerging two-dimensional micropatterning techniques offer powerful tools to construct controllable and well-organized microenvironment for single-cell level investigations with qualitative analysis,cellular standardization,and in vivo environment mimicking.Here,we provide an overview of the basic principle and characteristics of the two most widely-used micropatterning techniques,including photolithographic micropatterning and soft lithography micropatterning.Moreover,we summarize the application of micropatterning technique in controlling cytoskeleton,cell migration,nucleus and gene expression,as well as intercellular communication.展开更多
Collective cell migration is a coordinated movement of multi-cell systems essential for various processes throughout life.The collective motions often occur under spatial restrictions,hallmarked by the collective rota...Collective cell migration is a coordinated movement of multi-cell systems essential for various processes throughout life.The collective motions often occur under spatial restrictions,hallmarked by the collective rotation of epithelial cells confined in circular substrates.Here,we aim to explore how geometric shapes of confinement regulate this collective cell movement.We develop quantitative methods for cell velocity orientation analysis,and find that boundary cells exhibit stronger tangential ordering migration than inner cells in circular pattern.Furthermore,decreased tangential ordering movement capability of collective cells in triangular and square patterns are observed,due to the disturbance of cell motion at unsmooth corners of these patterns.On the other hand,the collective cell rotation is slightly affected by a convex defect of the circular pattern,while almost hindered with a concave defect,also resulting from different smoothness features of their boundaries.Numerical simulations employing cell Potts model well reproduce and extend experimental observations.Together,our results highlight the importance of boundary smoothness in the regulation of collective cell tangential ordering migration.展开更多
The transmembrane protein CD47,an innate immune checkpoint protein,plays a pivotal role in preventing healthy erythrocytes from immune clearance.Our study utilized stochastic optical reconstruction microscopy(STORM)an...The transmembrane protein CD47,an innate immune checkpoint protein,plays a pivotal role in preventing healthy erythrocytes from immune clearance.Our study utilized stochastic optical reconstruction microscopy(STORM)and single-molecule analysis to investigate the distribution of CD47 on the human erythrocyte membrane.Contrary to previous findings in mouse erythrocytes,we discovered that CD47 exists in randomly distributed monomers rather than in clusters across the human erythrocyte membrane.Using secondary antibody-induced crosslinking,we found that CD47 aggregates into stable clusters within minutes.By comparing these STORM results with those of the fully mobile protein CD59 and the cytoskeleton-bound membrane protein glycophorin C under similar conditions,as well as devising two-color STORM co-labeling and co-clustering experiments,we further quantitatively revealed an intermediate,self-limiting clustering behavior of CD47,elucidating its fractional(∼14%)attachment to the cytoskele-ton.Moreover,we report reductions in both the amount of CD47 and its clustering capability in aged erythrocytes,providing new insight into erythrocyte senescence.Together,the combination of STORM and secondary antibody-based crosslinking unveils the unique self-limiting clustering behavior of CD47 due to its fractional cytoskeleton attachment.展开更多
基金This work was supported by the Guangdong Major Project of Basic and Applied Basic Research(2020B0301030009)the National Key Research and Development Program of China(2022YFC3400600)National Natural Science Foundation of China(12174208,32227802,11874231,31801134 and 31870843)+2 种基金Tianjin Natural Science Foundation(20JCYBJC01010)China Postdoctoral Science Foundation(2020M680032)Fundamental Research Funds for the Central Universities(2122021337 and 2122021405).
文摘Actin cytoskeleton plays crucial roles in various cellular functions.Extracellular matrix(ECM)can modulate cell morphology by remodeling the internal cytoskeleton.To define how geometry of ECM regulates the organization of actin cytoskeleton,we plated individual NIH 3T3 cells on micropatterned substrates with distinct shapes and sizes.It was found that the stress fibers could form along the nonadhesive edges of T-shaped pattern,but were absent from the opening edge of V-shaped pattern,indicating that the organization of actin cytoskeleton was dependent on the mechanical environment.Furthermore,a secondary actin ring was observed on 50μm circular pattern while did not appear on 30μm and 40μm pattern,showing a size-dependent organization of actin cytoskeleton.Finally,osteoblasts,MDCK and A549 cells exhibited distinct organization of actin cytoskeleton on T-shaped pattern,suggesting a cell-type specificity in arrangement of actin cytoskeleton.Together,our findings brought novel insight into the organization of actin cytoskeleton on micropatterned environments.
基金the National Natural Science Foundation of China(Nos.11874231,11574165 and 31801134)Tianjin Natural Science Foundation(No.18JCQNJC02000)+1 种基金the PCSIRT(No.IRT 13R29)the 111 Project(No.B07013).
文摘As the indispensable oxygen-transporting cells,erythrocytes exhibit extreme deformability and amazing stability as they are subject to huge reversible shear stress and extrusion force during massive circulation in the body.The unique architecture of spectrin-actin-based membraneskeleton is considered to be responsible for such excellent mechanical properties of erythrocytes.Although erythrocytes have been recognized for more than 300 years,myriad questions about membrane-skeleton constantly attract people's attention.Here,we summarize the kinds of distinctive single-cell and single-molecule techniques that were used to investigate the structure and function of erythrocyte membrane-skeleton at macro and micro levels.
基金supported by the National Natural Science Foundation of China(Nos.12174208,32227802)National Key Research and Development Program of China(No.2022YFC3400600)+3 种基金Guangdong Major Project of Basic and Applied Basic Research(No.2020B0301030009)China Postdoctoral Science Foundation(No.2020 M680032)Fundamental Research Funds for the Central Universities(Nos.2122021337,2122021405)the 111 Project(No.B23045).
文摘Cells are highly sensitive to their geometrical and mechanical microenvironment that directly regulate cell shape,cytoskeleton and organelle,as well as the nucleus morphology and genetic expression.The emerging two-dimensional micropatterning techniques offer powerful tools to construct controllable and well-organized microenvironment for single-cell level investigations with qualitative analysis,cellular standardization,and in vivo environment mimicking.Here,we provide an overview of the basic principle and characteristics of the two most widely-used micropatterning techniques,including photolithographic micropatterning and soft lithography micropatterning.Moreover,we summarize the application of micropatterning technique in controlling cytoskeleton,cell migration,nucleus and gene expression,as well as intercellular communication.
基金supported by the National Natural Science Foundation of China(Nos.12174208 and 32227802)National Key Research and Development Program of China(No.2022YFC3400600)+2 种基金Guangdong Major Project of Basic and Applied Basic Research(No.2020B0301030009)Fundamental Research Funds for the Central Universities(Nos.2122021337 and 2122021405)the 111 Project(No.B23045).
文摘Collective cell migration is a coordinated movement of multi-cell systems essential for various processes throughout life.The collective motions often occur under spatial restrictions,hallmarked by the collective rotation of epithelial cells confined in circular substrates.Here,we aim to explore how geometric shapes of confinement regulate this collective cell movement.We develop quantitative methods for cell velocity orientation analysis,and find that boundary cells exhibit stronger tangential ordering migration than inner cells in circular pattern.Furthermore,decreased tangential ordering movement capability of collective cells in triangular and square patterns are observed,due to the disturbance of cell motion at unsmooth corners of these patterns.On the other hand,the collective cell rotation is slightly affected by a convex defect of the circular pattern,while almost hindered with a concave defect,also resulting from different smoothness features of their boundaries.Numerical simulations employing cell Potts model well reproduce and extend experimental observations.Together,our results highlight the importance of boundary smoothness in the regulation of collective cell tangential ordering migration.
基金supported by the National Key Research and Development Program of China(2022YFC3400600)the National Natural Science Foundation of China(11874231,32227802,and 12174208)+3 种基金Guangdong Major Project of Basic and Applied Basic Research(2020B0301030009)China Postdoctoral Science Foundation(2020M680032)the Fundamental Research Funds for the Central Universities(2122021337 and 2122021405)the Pew Charitable Trusts.
文摘The transmembrane protein CD47,an innate immune checkpoint protein,plays a pivotal role in preventing healthy erythrocytes from immune clearance.Our study utilized stochastic optical reconstruction microscopy(STORM)and single-molecule analysis to investigate the distribution of CD47 on the human erythrocyte membrane.Contrary to previous findings in mouse erythrocytes,we discovered that CD47 exists in randomly distributed monomers rather than in clusters across the human erythrocyte membrane.Using secondary antibody-induced crosslinking,we found that CD47 aggregates into stable clusters within minutes.By comparing these STORM results with those of the fully mobile protein CD59 and the cytoskeleton-bound membrane protein glycophorin C under similar conditions,as well as devising two-color STORM co-labeling and co-clustering experiments,we further quantitatively revealed an intermediate,self-limiting clustering behavior of CD47,elucidating its fractional(∼14%)attachment to the cytoskele-ton.Moreover,we report reductions in both the amount of CD47 and its clustering capability in aged erythrocytes,providing new insight into erythrocyte senescence.Together,the combination of STORM and secondary antibody-based crosslinking unveils the unique self-limiting clustering behavior of CD47 due to its fractional cytoskeleton attachment.
