Coupling intercellular adhesion, polarity and mechanical signals at epidermal junctions

细胞极化对正常发育、组织动态平衡和再生至关重要。对低等生物的研究确定了细胞和组织结构的重要调节因素。Par3-aPKC-Par6复合体中的极性蛋白介导不同的细胞过程，并将细胞形状的控制与调节生长、细胞命运和分化的信号通路结合起来。然而，这些保守的极性调节因子如何对哺乳动物的组织功能和疾病做出贡献还不是很清楚。皮肤是一个不断暴露在机械应力下的拉伸组织。极性蛋白是否以及如何驱动皮肤表皮等复层上皮细胞的极化，以及它们如何对该组织中的化学和机械信号起作用，在很大程度上尚不清楚。我们最近的工作揭示了Par3和aPKCl在皮肤癌中的重要作用，它们定位于连接部位，但不是胞浆Par3，促进了生长、生存和最终的肿瘤发生。我们进一步表明，小鼠表皮中Par3的失活会导致表皮屏障缺陷、干细胞衰退和过早分化。此外，Par3还调控P-钙粘附素的表面表达，从而调节表皮细胞与细胞之间的相互作用。在SPP1782中，我们将不同的生物物理方法与细胞生物学、遗传学和蛋白质生化方法相结合，揭示了意外的机制，通过这些机制，连接Par3将细胞间粘附处的机械信号与细胞反应结合起来，以确保表皮的动态平衡和屏障的完整性。我们破译了Par3促进Rho驱动的肌动球蛋白收缩能力，以促进细胞-细胞接触的内在力产生，这对维持紧密的连接屏障很重要。此外，我们的数据表明，极性蛋白在响应外界机械提示的动态连接重塑中起中介作用。有趣的是，纠正肌球蛋白的激活也足以挽救有丝分裂缺陷和Par3缺失时的异位分化，揭示了肌球蛋白调节在极性蛋白依赖的表皮自我更新和分化中的核心作用。在确定了连接蛋白PAR3对角质形成细胞力学的重要性后，第二个资助期的主要工作将致力于进一步解开细胞间粘连中的机械信号和极性信号之间的分子联系，这些信号介导细胞骨架和生化反应。我们将使用获得和丧失功能的方法、蛋白质组学、生物物理和成像方法来检查在Par3丢失后被解除调控的三类黏附分子的相关性。此外，我们将研究连接极性蛋白下游的机械力化学效应通路，并询问极性网络如何将机械信号传递到细胞内部，以指导菌株适应。总之，这个SPP1782项目将提供对细胞间连接、细胞极性和在重要的屏障形成上皮中力传递的耦合的分子洞察力。

Cell polarization is essential for proper development, tissue homeostasis and regeneration. Studies in lower organisms identified important regulators of cell and tissue architecture. Polarity proteins of the Par3-aPKC-Par6 complex mediate diverse cellular processes and couple control of cell shape to signalling pathways regulating growth, cell fate and differentiation. Yet, how these conserved polarity regulators contribute to mammalian tissue function and disease is much less clear. The skin is a tensile tissue constantly exposed to mechanical stress. Whether and how polarity proteins drive polarization in stratified epithelia like the skin epidermis and how they contribute to chemical and mechanical signalling in this tissue is largely unknown. Our recent work revealed important roles of Par3 and aPKCl in skin cancer, with junction-localized but not cytoplasmic Par3 promoting growth, survival and ultimately tumorigenesis. We further showed that Par3 inactivation in the mouse epidermis results in epidermal barrier defects, stem cell decline and premature differentiation. Moreover, Par3 controls P-cadherin surface expression, thereby regulating epidermal cell-cell interactions. Within the SPP1782 we combined different biophysical methods with cell biological, genetic and proteinbiochemical approaches to reveal unexpected mechanisms through which junctional Par3 couples mechanical signals at intercellular adhesions with cellular responses to ensure epidermal homeostasis and barrier integrity. We deciphered that Par3 promotes Rho-driven actomyosin contractility to foster intrinsic force generation at cell-cell contacts, important to maintain the tight junctional barrier. Moreover, our data indicate that polarity proteins mediate dynamic junction remodelling in response to external mechanical cues. Intriguingly, correcting myosin activation was also sufficient to rescue mitotic defects and ectopic differentiation upon Par3 loss, unravelling a central role of actomyosin regulation in polarity protein-dependent epidermal self-renewal and differentiation. Having established the importance of junctional Par3 for keratinocyte mechanics, key efforts in the second funding period will aim to unravel further molecular links between mechanical and polarity signalling at intercellular adhesions that mediate cytoskeletal and biochemical responses. We will use gain- and loss-of-function approaches, proteomics, biophysical and imaging methods to examine the relevance of three classes of adhesion molecules that are deregulated following Par3 loss. Moreover, we will investigate mechanochemical effector pathways downstream of junctional polarity proteins and ask how polarity networks transmit mechanical cues to the cell’s interior to instruct strain adaptation. Together, this SPP1782 project will provide molecular insight into the coupling of intercellular junctions, cell polarity and force transmission in an important barrier-forming epithelium.