A role of plakophilin 3 in keratinocyte proliferation

plakophilin 3 在角质形成细胞增殖中的作用

• 批准号: 273121986
• 负责人: Professorin Dr. Mechthild Hatzfeld
• 金额: --
• 依托单位: Arbeitsgruppe Pathobiochemie
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/273121986?language=en
• 关键词: role; plakophilin; keratinocyte; proliferation

Desmosomes are intercellular adhesive contacts that are especially abundant in tissues prone to mechanical strain such as the skin and the heart. Our data suggest that members of the plakophilin family of desmosomal proteins control desmosome adhesive strength, size and number. Whereas plakophilin 1 strengthens intercellular adhesion and increases desmosome size in vivo and in vitro, plakophilin 3 renders desmosomes more dynamic. In agreement, our mouse knockout studies reveal severe skin fragility in plakophilin 1 KO mice with postnatal lethality. Moreover, we have shown that plakophilins 1 and 3 act as switches between desmosomal adhesion, cell proliferation, cell migration and anchorage independent growth. Cytoplasmic plakophilin 1 stimulates proliferation by associating with the translation initiation complex to increase protein biosynthesis. Activation of these regulatory functions depends on plakophilin 1 phosphorylation by the Akt2 kinase downstream of IGF1/insulin signaling. Phosphorylated plakophilin 1 becomes trapped in the cytoplasm by 14-3-3 which interferes with its incorporation into desmosomes. We found that plakophilin 3 knockout keratinocytes reveal significantly reduced proliferation rates, in agreement with growth retardation of plakophilin 3 KO mice, whereas its overexpression promotes proliferation of keratinocytes. Our comparative gene expression analysis revealed that 30% of all genes downregulated in plakophilin 3 KO cells are cell cycle related, many of them being E2F targets. Moreover, we find that plakophilin 3 translocates to the mitotic spindle. The identification of several putative interacting proteins essential for mitotic spindle formation and dynamics suggests an additional role in cell cycle progression. Based on these data we propose the hypothesis that plakophilin 3 is involved in cell cycle control by (a) regulating S phase entry via control of E2F/Rb activity and (b) regulating the mitotic spindle in association with microtubule binding proteins. Our project aims to verify this hypothesis by validation of E2F target gene regulation and an analysis of the molecular mechanism by which plakophilin 3 might influence E2F/Rb activity to promote S-phase entry. Moreover, plakophilin 3 localization and protein interactions during mitosis and mechanisms controlling its changes in localization and protein interactions in a cell cycle dependent manner will be studied.We expect that our studies will have a major impact on understanding the interplay between desmosomal adhesion and proliferation which has to be tightly balanced to guarantee epidermal homeostasis. Moreover, we will gain insight into the role of plakophilin 3 in modulating desmosomes to allow for mobility without interfering with cohesion during mitosis.

桥粒是一种细胞间的粘连接触，在皮肤和心脏等易受机械应变的组织中尤其丰富。我们的数据表明，桥粒蛋白中的亲核蛋白家族成员控制着桥粒的黏附强度、大小和数量。在体内和体外，亲板素1增强细胞间的黏附并增加桥粒大小，而亲板素3使桥粒更具活力。一致的是，我们的小鼠基因敲除研究显示，具有出生后致死性的PLAK-1KO小鼠皮肤严重脆弱。此外，我们还发现，在桥粒黏附、细胞增殖、细胞迁移和锚定非依赖性生长之间，Plakopilin1和Plakopilin3起着开关作用。细胞质中的蛋白亲和素1通过与翻译起始复合体结合，促进蛋白质生物合成，从而刺激细胞增殖。这些调节功能的激活依赖于IGF1/胰岛素信号通路下游Akt2激酶对Plakophin1的磷酸化。通过14-3-3，磷酸化的蛋白亲和素1被困在细胞质中，这干扰了它与桥粒的整合。我们发现，PK基因敲除的角质形成细胞的增殖率显著降低，这与PK-3KO小鼠的生长迟缓一致，而其过表达则促进角质形成细胞的增殖。我们的比较基因表达分析表明，在PLAKAPILIN3KO细胞中下调的所有基因中有30%与细胞周期有关，其中许多是E2F靶标。此外，我们还发现亲斑蛋白3易位到有丝分裂纺锤体。对有丝分裂纺锤体形成和动力学所必需的几种可能的相互作用蛋白的鉴定表明，在细胞周期进程中还有另外的作用。在这些数据的基础上，我们提出了亲膜蛋白3参与细胞周期调控的假说：(A)通过调控E2F/Rb活性来调节S的时相进入；(B)通过微管结合蛋白调节有丝分裂纺锤体。我们的项目旨在通过验证E2F靶基因的调控和分析PPA3可能影响E2F/Rb活性促进S进入时相的分子机制来验证这一假说。此外，我们还将研究有丝分裂过程中PLAKAPRIN 3的定位和蛋白质相互作用，以及以细胞周期依赖的方式控制其定位变化和蛋白质相互作用的机制。我们预计，我们的研究将对理解桥粒黏附和增殖之间的相互作用产生重要影响，桥粒黏附和增殖之间的相互作用必须保持紧密平衡，以确保表皮的动态平衡。此外，我们还将深入了解在有丝分裂过程中，亲膜蛋白3在调节桥粒的作用，从而在不干扰凝聚力的情况下实现移动性。