Inner nuclear membrane protein TMEM201 maintains endothelial cell migration and angiogenesis by interacting with the LINC complex

The nuclear envelope comprises the outer nuclear membrane,inner nuclear membrane(INM),and nucleopore.Although∼60 INM proteins have been identified,only a few of them have been well characterized,revealing their crucial roles.Our group focused on the INM protein transmembrane protein 201(TMEM201),whose role in cellular function remains to be defined.In this study,we investigated the role of TMEM201 in endothelial cell migration and angiogenesis.Depletion of TMEM201 expression by short hairpin RNA-mediated interference impeded human umbilical vein endothelial cell(HUVEC)angiogenic behavior in tube formation and fibrin gel bead sprouting assays.Meanwhile,TMEM201-deficient HUVECs exhibited impaired migration ability.We next explored the underlying mechanism and found that the N-terminal of TMEM201 interacted with the linker of nucleoskeleton and cytoskeleton complex and was required for regulating endothelial cell migration and angiogenesis.These in vitro findings were further confirmed by using in vivo models.In Tmem201-knockout mice,retinal vessel development was arrested and aortic ring sprouting was defective.In addition,loss of tmem201 impaired zebrafish intersegmental vessel development.In summary,TMEM201 was shown to regulate endothelial cell migration and control the process of angiogenesis.This study is the first to reveal the role of INM proteins in the vascular system and angiogenesis.

LINC复合体与细胞内机械力传导

细胞作为力学感受器,可以感知、传递施加在其表面的机械力并调整自身力学性能,维持自身稳定。机械力从细胞表面或细胞质传递到细胞核依赖于完整的细胞骨架系统,该系统包括细胞质骨架和细胞核骨架两部分,而LINC复合体则是两者实现机械连接的桥梁,因此其在细胞内机械力传导中发挥着重要作用。综述LINC复合体传导机械力的结构基础和机械力传导引起的细胞核形态、转录因子的出入核以及染色质的空间构象的改变,为进一步探讨LINC复合体在细胞机械力传导过程中的作用及其对基因表达的影响奠定基础。

Do tensile and shear forces exerted on cells influence mechanotransduction through stored energy considerations?

All tissues in the body are subjected externally to gravity and internally by collagenfibril and cellular retractive forces that create stress and energy equilibrium required for homeostasis.Mechanotransduction involves mechanical work(force through a distance)and energy storage as kinetic and potential energy.This leads to changes in cell mitosis or apoptosis and the synthesis or loss of tissue components.It involves the application of energy directly to cells through integrin-mediated processes,cell-cell connections,stretching of the cell cytoplasm,and activation of the cell nucleus via yes-associated protein(YAP)and transcriptional coactivator with PDZ-motif(TAZ).These processes involve numerous complexes,intermediate molecules,and multiple pathways.Several pathways have been identified from research studies on vertebrate cell culture and from studies in invertebrates.These pathways involve mechanosensors and other molecules that activate the pathways.This review discusses the mitogen-activated protein kinase(MAPK)family,Hippo,Hedgehog,and Wingless-related integration site(WNT)/βcatenin signaling pathways.The mediators covered includeβcatenin,ion channels,growth factors,hormone receptors,members of the Ras superfamily,and components of the linker of nucleoskeleton and cytoskeleton(LINC)complex.However,the interrelationship among the different pathways remains to be clarified.Integrin-mediated mechanotransduction involves direct tensile loading and energy applied to the cell membrane via collagenfibril stretching.This energy is transferred between cells by stretching the cell-cell connections involving cadherins and the WNT/βcatenin pathway.These alterations induce changes in intracellular events in the cytoskeleton and nuclear skeleton caused by the release of YAP and TAZ.These coactivators then penetrate through the nuclear pores and influence nuclear cell function.Alteration in the balance of forces and energy applied to cells and tissues is hypothesized to shift the cell-extracellular matrix mechanical equilibrium by modifying mechanotransduction.The shift in equilibrium can lead to either tissue synthesis,genetic modifications,or promotefibrotic diseases,including epithelial cell-derived cancers,depending on the local metabolic conditions.

人睾丸蛋白SPAG4L与KASH结构域蛋白Nesprin-3的相互作用

目的探讨含SUN(Sad,UNC-84)结构域的人睾丸蛋白SPAG4L(sperm-associated antigen 4 like)与具有KASH(Klarsicht,ANC-1 and Syne homology)结构域的核膜血影重复蛋白3(nuclear envelop spectrin repeat proteins 3,Nesprin-3)之间的相互作用。方法用生物信息学方法对SPAG4L蛋白进行分析,通过体外转染实验,观察SPAG4L的亚细胞定位;并用免疫荧光技术、免疫共沉淀和双分子荧光互补实验检测SPAG4L是否与KASH结构域蛋白Nesprin-3存在相互作用。结果生物信息学分析结果表明,SPAG4L蛋白具有跨膜结构;亚细胞定位结果发现,SPAG4L蛋白定位于核膜和胞浆;免疫荧光、免疫共沉淀和双分子荧光互补实验结果表明,SPAG4L蛋白与Nesprin-3蛋白质相互作用,形成LINC(linkers of the nucleoskeleton to the cytoskeleton)复合物。结论 SPAG4L与Nesprin-3能够相互作用,形成LINC复合物,对了解SPAG4L蛋白在精子发生中的作用具有重要的意义。

海鞘细胞核骨架与细胞质骨架连接复合体相关基因的克隆及表达分析

为了研究LINC(Linker of nucleoskeleton and cytoskeleton)复合体相关基因KASH(Klarsicht/Anc/Syne-1)和SUN(Sad1/UNC84)在海鞘(Ciona robusta)细胞核位置决定中的角色和作用,本研究鉴定并分别克隆了海鞘LINC复合体中编码2种Cr_KASH蛋白和2种Cr_SUN蛋白的基因。荧光定量PCR方法分析了上述4个基因在海鞘胚胎各个时期的表达量,发现在胚胎发育的尾芽延伸期,几种基因的表达量均有不同程度的提高。原位杂交方法揭示Cr_KASH1、Cr_SUN1和Cr_SUN2特异表达在海鞘胚胎的脊索组织中,而Cr_KASH2则特异表达在肌肉组织。在NIH/3T3细胞中的亚细胞定位分析证实,上述LINC复合体蛋白与不同的细胞骨架交联。通过破坏细胞骨架微丝和微管蛋白的聚合发现会影响KASH和SUN在细胞中的定位和结构,暗示海鞘LINC复合体成分和细胞质骨架蛋白协同作用调控细胞核的形态和位置。本文研究结果表明,在海鞘不同胚胎发育阶段和不同组织中,细胞核骨架或许通过不同LINC蛋白组分与细胞骨架结合影响细胞核的行为。