Cell biology of osteoclasts: GTPases, exchange factors and actin dynamics during bone degradation

破骨细胞是专门用于骨降解的多核细胞。为了消化骨骼，它们需要附着在骨骼表面，通过将富含肌动蛋白的足体重新组织成紧密带来建立密封区，并修改膜运输的关键步骤。我们的建议集中在FGD6，一种在成骨细胞形成过程中强烈上调的Rho鸟嘌呤交换因子。我们的初步结果表明，FGD6调节足体的形成，并潜在地调节后体带周围的内体的定位。为了了解FGD6的功能，有必要1)实时研究其动态行为，2)仔细阐明内源性FGD6缺失的破骨细胞的表型，3)阐明FGD6的PH和FYVE磷酸肌醇结合结构域对其功能的贡献4)确定FGD6的结合伙伴，特别是激活的RhoGTPase，以及5)了解FGD6的激活和/或定位如何受到Src和PI-3激酶信号的影响。我们想要验证的假设是，FGD6通过其PH结构域与质膜PI(3，4，5)P3之间的相互作用以及通过其FYVE结构域与早期内体的PI3-P之间的相互作用控制足体动力学，这一过程可能受到Src依赖的磷酸化的调节。这些研究应该为其他RhoGEF的功能探索铺平道路，这些RhoGEF与FGD6一样，在破骨细胞形成过程中强烈上调。

Osteoclasts are multinucleated cells specialized in bone degradation. To digest bone, they need to adhere onto its surface, to build a sealing zone by reorganizing their actin-rich podosomes into tight belts and to modify key steps of membrane traffic. Our proposal focuses on FGD6, a Rho guanine exchange factor strongly upregulated during osteolastogenesis. Our preliminary results indicate that FGD6 regulates podosome formation and potentially the localization of endosomes around podosomal belts. To understand FGD6 function, it will be necessary 1) to study its dynamic behavior in real time, 2) to carefully illustrate the phenotype of osteoclasts depleted in endogenous FGD6, 3) to illustrate how the PH and FYVE phosphoinositide binding domains of FGD6 contribute to its function 4) to identify the FGD6 binding partners, in particular the activated RhoGTPase, and 5) to understand how FGD6 activation and/or localization can be affected by Src and PI-3 kinase signaling. The hypothesis that we want to test is that FGD6 controls podosome dynamics via interactions between its PH domains and PI(3,4,5)P3 of the plasma membrane as well as the localization of early endosomes around podosomal belts via interactions between its FYVE domain and PI3-P of early endosomes, processes that might be regulated by Src-dependent phosphorylation. These studies should pave the way for the functional exploration of other RhoGEFs, which like FGD6 are strongly upregulated during osteoclastogenesis.