Dynamic Regulation of Erythropoietin Gene Expression in Mammals.

贫血是一种常见的医学病症，与显著的发病率和死亡率相关， 特别是如果存在其他疾病如心力衰竭。重组技术的发展 促红细胞生成素（Epo）是一种在成人贫血时由肾脏和肝脏产生的促红细胞激素， 彻底改变了贫血治疗方法不幸的是，非生理性推注Epo给药也 促进血栓形成、高血压和可能的癌症生长。这部分是因为 外源性Epo的施用缺乏内源性Epo的正常反馈调节特征， 其甚至在红细胞增多明显之前就被迅速下调。内源性Epo产生和 铁的吸收是由应激反应的异二聚体转录因子低氧严格控制的 诱导因子2（HIF-2），其作用受氧依赖性和氧非依赖性调节。 翻译后修饰（PTM）。氧依赖性PTM包括脯氨酰的羟基化 氧依赖性脯氨酰介导的HIF α蛋白特定元件中的残基 羟化酶（PHD），一个三种相关蛋白的家族，其表现出对HIF-1和 HIF-2 α亚单位。 PHD抑制剂（PHDi）的开发最近在经过协调一致的研究后进入市场。 虽然这些药物的研发工作已经进行了近二十年，但其长期安全性仍然未知。的 PHDi的作用机制涉及稳定和/或增强HIF-2信号传导的活性。 然而，HIF-1信号可能也被激活，因为缺乏对PHDi的完全特异性。 以及PHD蛋白对HIF本身的干扰。虽然PHDi可能代表一个 与外源性Epo推注相比，PHDi治疗仍必须严格控制 因为它们也缺乏与控制内源性Epo产生相关的反馈行为。 此外，由于这些药剂最近才进入市场， 在世界上的许多地区，与它们在真实的世界环境中使用相关的副作用现在才开始出现。 光有病例报告称，使用这些药物会增加血栓形成， 考虑到HIF-2的长期增强， 肠道中的信号传导也可以刺激铁的吸收和利用。 为了利用Epo表达的正常反馈调节特征，我们必须 了解HIF-2信号在缺氧过程中如何在动态和时间意义上发生。一 增强HIF-2信号传导的机制涉及循环乙酰化和去乙酰化后， 翻译修饰该方法中的限速步骤是提供乙酰辅酶A用于 HIF-2的乙酰化，其由乙酸依赖性乙酰辅酶A产生器酰基辅酶A产生 合成酶2（Acss 2）。acss 2在缺氧的早期阶段是活跃的，当细胞在缺氧时产生乙酸盐。 应对这种环境压力。在这个建议中，我们将定义周围的相互作用组， Acss 2和HIF-2在常氧条件下以及在缺氧早期和晚期。我们将 还确定是否合理的药物治疗方法，基于对HIF-2如何 乙酰化/脱乙酰化完成后，可用于治疗慢性贫血相关的小鼠 患有慢性肾病

Anemia is a common medical condition associated with significant morbidity and mortality, especially if present with other diseases such as heart failure. The development of recombinant Erythropoietin (Epo), a pro-erythrocyte hormone produced in adult kidney and liver during anemia, revolutionized anemia treatment. Unfortunately, non-physiological bolus Epo administration also promotes thrombosis, hypertension, and possibly cancer growth. This occurs in part because administration of exogenous Epo lacks the normal feedback regulatory features of endogenous Epo, which is rapidly down-regulated even before erythrocytosis is evident. Endogenous Epo production and iron uptake are tightly controlled by the stress-responsive, heterodimeric transcription factor Hypoxia Inducible Factor 2 (HIF-2), whose actions are regulated by oxygen-dependent and oxygen-independent post-translational modifications (PTM). The oxygen-dependent PTM include hydroxylation of prolyl residues in specific elements of the HIF alpha proteins mediated by oxygen-dependent prolyl hydroxylases (PHD), a family of three related proteins that exhibit differential recognition of HIF-1 and HIF-2 alpha subunits. Development of PHD inhibitors (PHDi) have recently come to market after a concerted development effort over nearly two decades, but their long-term safety remains unknown. The mechanism of action for PHDi involves stabilization and/or enhanced activity of HIF-2 signaling. However, HIF-1 signaling is likely activated as well given the lack of complete specificity for the PHDi as well as cross-talk that PHD proteins have for HIF themselves. Although PHDi may represent an improvement over exogenous bolus Epo administration, PHDi treatment must still be tightly controlled as they also lack feedback behaviors associated with control of endogenous Epo production. Furthermore, because these agents have only recently come to the market and are being used in select regions of the world, side effects associated with their use in real world settings are only now coming to light. There have been case reports of increased thrombosis with their use and there is at least a theoretical concern for development of iron overload, given that prolonged augmentation of HIF-2 signaling in the gut may also stimulate iron uptake and utilization. In order to leverage the normal feedback regulatory features of Epo expression, we must understand how HIF-2 signaling occurs in a dynamic and temporal sense during hypoxia. One mechanism for enhancing HIF-2 signaling involves cyclical acetylation and deacetylation post- translational modifications. A rate-limiting step in this process is provision of acetyl CoA for use in acetylation of HIF-2, which is generated by the acetate-dependent acetyl CoA generator, acyl CoA synthetase 2 (Acss2). Acss2 is active in the early phase of hypoxia when cells generate acetate in response to this environmental stress. In this proposal, we will define the interactome surrounding Acss2 and HIF-2 under normoxic conditions as well as under early and late hypoxia periods. We will also determine if a rational drug therapy approach, based upon the knowledge of how HIF-2 acetylation/deacetylation is accomplished, can be used to treat mice with chronic anemia associated with chronic kidney disease.