Calcium sensing receptor and scaffolds

描述（由申请人提供）：钙感应受体（CaR）将细胞外钙和代谢物（抗tino酸、多胺）的局部变化转导为细胞内信号，包括细胞代谢和分泌的急性改变，以及细胞增殖和分化的长期变化。CaR通过控制甲状旁腺PTH分泌和肾脏钙排泄/吸收平衡，对维持全身钙水平至关重要。CaR信号传导的细胞特异性变异性可能部分是由于CaR与支架蛋白的相互作用和细胞特异性信号传导复合物的形成。细胞骨架蛋白丝蛋白A是CaR的支架，是CaR介导的MARK信号激活和减缓CaR降解所必需的。初步数据表明，CaR与抑制因子-1和-2的相互作用可调节细胞内钙反应和MARK信号。为了理解蛋白质支架如何调节CaR介导的信号传导，我们将检验以下假设：(1)状态依赖性阻滞蛋白-1结合是否调节CaR反应性？(2) car介导的MARK信号通路是否由丝蛋白A和阻滞蛋白2组织，该信号通路是否包括EGF受体的反激活？(3)丝蛋白A是否通过保护CaR免受蛋白酶体介导的降解而起到稳定细胞内CaR水平的分子护送作用？我们将结合生物化学方法与细胞内钙和/或共聚焦成像，使用人类CaR和突变和缺失结构。所提出的实验将增加我们对蛋白质支架阻滞蛋白-1和-2和丝蛋白A在增强CaR信号传导特异性中的作用的理解。人类CaR中有超过40个突变与疾病有关，彻底改变了甲状旁腺疾病的临床解剖。CaR变构调节剂在治疗原发性甲状旁腺功能亢进和低下、肾脏疾病引起的继发性甲状旁腺功能改变和骨质疏松症方面具有临床应用价值。CaR刺激细胞增殖和PTHrP分泌，提示CaR在恶性肿瘤的病理生理后果中起着增强作用。CaR表达和/或急性或长期信号的失调部分可能是与支架蛋白相互作用改变的结果，支架蛋白增强了CaR信号的特异性并调节了CaR的稳定性。了解支架在CaR功能中的作用可能为治疗钙处理性疾病的药物干预提供新的、特定的位点。

DESCRIPTION (provided by applicant): The calcium sensing receptor (CaR) transduces local changes in extracellular calcium and metabolites (anrtino acids, polyamines) into intracellular signals, which include acute alterations in cell metabolism and secretion, and long term changes in cell proliferation and differentiation. CaR is critical to maintenance of systemic calcium levels, by controlling parathyroid PTH secretion and the balance of renal calcium excretion/resorption. Cell-specific variability in CaR signaling may in part be due to interaction(s) of CaR with scaffold proteins and the formation of cell-specific signaling complexes. The cytoskeletal protein filamin A is a scaffold for CaR, required for CaR-mediated activation of MARK signaling, and slowing CaR degradation. Preliminary data suggests CaR interactions with arrestins-1 and -2 modulate intracellular calcium responses and MARK signaling. To understand how protein scaffolds regulate CaR-mediated signaling, we will test the following hypotheses: (1) Does state-dependent arrestin-1 binding regulate CaR responsiveness? (2) Is CaR-mediated MARK signaling organized by filamin A and arrestin-2, and does the signaling pathway include transactivation of the EGF receptor? (3) Does filamin A act as a molecular escort to stabilize cellular CaR levels by protecting CaR from proteasome-mediated degradation? We will combine biochemical approaches with intracellular calcium and/or confocal imaging, using human CaR and mutant and deletion constructs. The proposed experiments will increase our understanding of the role of the protein scaffolds arrestin-1 and -2 and filamin A in enhancing the specificity of CaR signaling. More than 40 mutations in human CaR have been linked to disease, revolutionizing the clinical dissection of parathyroidisms. Allosteric modulators of CaR have clinical utility for treatment of primary hyper- and hypo- parathyroidism, secondary changes in parathyroid function resulting from renal disease, and osteoporosis. CaR stimulates cell proliferation as well as PTHrP secretion, suggesting role for CaR in potentiating the pathophysiological consequences of malignancies. Dysregulation of CaR expression and/or acute or long term signaling may, in part, be the result of altered interactions with scaffold proteins which serve to enhance CaR signaling specificity and to regulate CaR stability. Understand the role of scaffolds in CaR function may provide novel, specific sites for pharmacological intervention in treatment of calcium handling diseases.