Mechanical regulation of cytoskeleton guides beta-catenin effect on MSC fate

细胞骨架的机械调节引导β-连环蛋白对MSC命运的影响

• 批准号: 9252230
• 负责人: Janet E Rubin
• 金额: $ 33.44万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9252230
• 关键词: AKT inhibition; Actins; Adhesions; Adipocytes; Adipose tissue; Adult; Bone Formation Stimulation; Bone Marrow; Cell Lineage; Cell Nucleus; Cells; Complex; Cytoskeletal Modeling; Cytoskeleton; Data; Exercise; F-Actin; Fatty acid glycerol esters; Focal Adhesions; Goals; Grant; Health; Histologic; In Vitro; Knock-out; Knockout Mice; Lamins; Marrow; Measures; Mechanics; Mesenchymal Stem Cells; Methods; Mus; Nuclear; Obesity; Osteogenesis; Pathway interactions; Periodicity; Pharmacology; Phosphotransferases; Physicians; Physiological; Population; Proteins; Recruitment Activity; Regulation; Regulatory Pathway; Resistance; Role; Running; Scientist; Signal Pathway; Signal Transduction; Stem cells; Stress Fibers; Structure; Tamoxifen; Testing; Time; Work; base; beta catenin; bone; bone cell; bone fragility; bone health; bone strength; improved; in vivo; innovation; lipid biosynthesis; mechanical force; mechanical load; mouse model; osteogenic; osteoprogenitor cell; prevent; public health relevance; response; sedentary; skeletal; synergism; trafficking

 DESCRIPTION (provided by applicant): Exercise builds skeletal strength by inducing new bone formation, which is accompanied by decreased marrow adipose tissue (MAT). A lack of exercise enhances MSC adipogenesis, and increased MAT has been shown to be associated with bone fragility. Understanding mechanisms by which exercise regulates mdMSC lineage allocation to increase osteogenesis and decrease adipogenesis is thus significant for managing skeletal health in an increasingly sedentary population. We have shown that mechanical activation and nuclear trafficking of ßcatenin prevents adipogenesis and promotes MSC entry into osteoprogenitor lineage. ßcatenin signaling is initiated by recruitment of Fyn kinase to foca adhesions during strain, followed by Fyn activation of the mTORC2->Akt->GSK3ß inhibition cascade leading to increased ßcatenin. ßcatenin effects depend on cellular context: in mdMSC, increased cytoskeletal structure improves osteogenesis and cytoskeletal deficiency promotes adipogenesis. The Fyn/mTORC2 proximal signal also activates RhoA to build complexity and connectivity of the cytoskeleton. We propose that mechanically induced cytoskeletal reorganization promotes nuclear entry of ßcatenin, underwriting its anti-adipogenic, pro-osteogenic effect. Importantly, the role of focal adhesion-based Fyn in initiating ßcatenin and RhoA effectors may be unique to the mdMSC, potentially providing a specific pharmacologic target to enhance MSC response to exercise. To test this, we will delineate the synergistic contributions of Fyn/mTORC2's two downstream effectors, RhoA and ßcatenin, to MSC lineage fate allocation in vitro and in vivo. We will find if RhoA generated cytoskeletal reorganization is necessary or sufficient to alter mdMSC cell fate and define how mechanical load activates RhoA (SA1). In SA2, we ask if activation of ßcatenin can alter cell fate in the face of cytoskeletal deficiency, inquiring whether the cytoskeleton drives nuclear/cytoplasmic partitioning of ßcatenin and considering if the actin-nesprin-LINC-nuclear tether might guide ßcatenin into the nucleus. In SA3, using an innovative method to quantify and localize MAT, combined with measures of bone formation, we will investigate mechanical control of mdMSC lineage in vivo, asking if the Fyn/mTORC2 signaling unique to mdMSC is critical during exercise stimulation of bone formation; for this we will generate a Tamoxifen driven Prx1-Cre knock out of Fyn.

 描述（由申请人提供）：运动通过诱导新骨形成来增强骨骼强度，同时伴随着骨髓脂肪组织（MAT）的减少。缺乏运动会增强间充质干细胞的脂肪生成，而 MAT 的增加已被证明与骨脆性有关。因此，了解运动调节 mdMSC 谱系分配以增加成骨和减少脂肪生成的机制对于管理日益久坐人群的骨骼健康具有重要意义。我们已经证明，β连环蛋白的机械激活和核运输可防止脂肪生成并促进 MSC 进入骨祖细胞谱系。 ßcatenin 信号传导是通过在应变期间将 Fyn 激酶募集到粘着灶而启动的，随后 Fyn 激活 mTORC2->Akt->GSK3ß 抑制级联，导致 ßcatenin 增加。 β连环蛋白的作用取决于细胞环境：在 mdMSC 中，细胞骨架结构的增加可改善成骨作用，而细胞骨架的缺乏可促进脂肪形成。 Fyn/mTORC2 近端信号还激活 RhoA 以构建细胞骨架的复杂性和连接性。我们认为机械诱导的细胞骨架重组促进β连环蛋白进入核，从而保证其抗脂肪形成、促骨形成作用。重要的是，基于粘着斑的 Fyn 在启动 ßcatenin 和 RhoA 效应器中的作用可能是 mdMSC 所独有的，可能提供特定的药理学靶点来增强 MSC 对运动的反应。为了测试这一点，我们将描述 Fyn/mTORC2 的两个下游效应器 RhoA 和 ßcatenin 对体外和体内 MSC 谱系命运分配的协同贡献。我们会发现 RhoA 产生的细胞骨架重组是否是 对于改变 mdMSC 细胞命运并定义机械负荷如何激活 RhoA (SA1) 是必要或充分的。在 SA2 中，我们询问 ßcatenin 的激活是否可以在细胞骨架缺陷时改变细胞命运，探究细胞骨架是否驱动 ßcatenin 的核/细胞质分配 并考虑肌动蛋白-nesprin-LINC-核系链是否可以引导 ßcatenin 进入细胞核。在SA3中，我们将使用创新方法量化和定位MAT，结合骨形成的测量，研究体内mdMSC谱系的机械控制，询问mdMSC特有的Fyn/mTORC2信号在运动刺激骨形成过程中是否至关重要；为此，我们将产生他莫昔芬驱动的 Prx1-Cre 敲除 Fyn。