NBCe1-mediated Regulation of HCO3- is a Novel Mechanism Underlying Metabolic Reprogramming and Cystogenesis

项目总结/摘要 葡萄糖刺激的胰岛素分泌（GSIS）的丧失是一个重要的病理生理事件， 2型糖尿病（T2 DM）中高血糖症的发展。最近的证据表明， 糖尿病中的GSIS与线粒体功能降低的代谢重编程有关;然而， 这些现象背后的机制在很大程度上仍然未知。单细胞转录组学研究进展 的人β细胞将SLC 4A 4鉴定为在T2 DM β细胞中高度表达的少数独特基因之一， 在非糖尿病β细胞中被抑制。Slc 4a 4编码胰腺中的Na+-nHCO 3-协同转运蛋白NBCe 1B， 在调节细胞内pH（pHi）中起关键作用。重要的是，NBCe 1的激活增加已经被证实是一种新的生物学效应。 与增强的细胞内糖酵解和受损的线粒体功能有关，这表明它可能 导致GSIS丢失和随后的T2 DM发展。初步的论文研究支持这一点 假设并证明抑制β细胞中NBCe 1B活性改善体外GSIS并增强 体内葡萄糖耐量。这些累积的观察使我们开发了一个博士论文方向 总体目标是表征NBCe 1B作为β细胞代谢的新型调节剂的作用， 2型糖尿病的功能障碍。因此，特定目的1（F99）将检验T2 DM中β细胞功能障碍的假设， 是由代谢重编程驱动的，代谢重编程由NBCe 1B激活的细胞碱化介导。给定 NBCe 1在维持全身pH稳态中的关键作用，F99独特地定位我来阐明 与K 00期肾脏酸碱平衡失调相关的新机制。 具体地说，NBCe 1的A-亚型（NBCe 1A）是HCO 3重吸收的关键机制， 肾NBCe 1A缺失与集合管内代谢性酸中毒和皮质囊肿相关 （CD）。可溶性腺苷酸环化酶（sAC）已被确定为CD内的HCO 3-传感器。以前的工作 表明受损的NBCe 1A介导的HCO 3重吸收激活sAC-cAMP/PKA介导的 发信号。有趣的是，CD内持续的cAMP/PKA激活也被证明是一个关键因素。 多囊肾病（PKD）的发病机制因此，主 我提出的博士后研究方向的目标是将NBCe 1A的作用描述为一种新的 通过激活sAC-cAMP/PKA信号通路调节囊形成。具体目标2 (K00)将测试这一假设，即受损的NBCe 1A介导的HCO 3重吸收激活可溶性腺苷酸 集合管中环化酶-cAMP/PKA信号级联促进模型中的增殖和囊肿形成 的PKD。F99和K 00将共同推动我实现我的长期目标，领导一个独立的 肾脏病学的研究项目

PROJECT SUMMARY/ABSTRACT The loss of glucose-stimulated insulin secretion (GSIS) is a critical pathophysiological event precipitating development of hyperglycemia in Type 2 diabetes mellitus (T2DM). Recent evidence suggests that loss of GSIS in diabetes is associated with metabolic reprogramming toward reduced mitochondrial function; however mechanisms underlying these observations remain largely unknown. Recent single cell transcriptomics studies of human β-cells identified SLC4A4 as one of few unique genes highly expressed in T2DM β-cells and repressed in non-diabetic β-cells. Slc4a4 encodes for Na+-nHCO3- cotransporter, NBCe1B in the pancreas and plays a key role in regulating intracellular pH (pHi). Importantly, increased activation of NBCe1 has been associated with enhanced intracellular glycolysis and impaired mitochondrial function suggesting it may contribute to loss of GSIS and consequent development of T2DM. Preliminary dissertation studies support this hypothesis and demonstrate that inhibition of NBCe1B activity in β-cells improves GSIS in vitro and enhances glucose tolerance in vivo. These cumulative observations led us to develop a doctoral dissertation direction with an overall objective to characterize the role of NBCe1B as a novel regulator of β-cell metabolism and dysfunction in T2DM. Accordingly, Specific Aim 1 (F99) will test the hypothesis that β-cell dysfunction in T2DM is driven by metabolic reprogramming mediated by cellular alkalization through activation of NBCe1B. Given the critical role of NBCe1 in maintaining systemic pH homeostasis, the F99 uniquely positions me to elucidate novel mechanisms associated with dysregulation of acid-base balance in the kidney during the K00 phase. Specifically, the A-isoform of NBCe1 (NBCe1A) functions as the key mechanism of HCO3- reabsorption in the kidney. Deletion of NBCe1A is associated with metabolic acidosis and cortical cysts within the collecting duct (CD). Soluble adenylyl cyclase (sAC) has been identified as a HCO3- sensor within the CD. Previous work demonstrated that impaired NBCe1A-mediated HCO3- reabsorption activates sAC-cAMP/PKA mediated signaling. Interestingly, persistent cAMP/PKA activation within the CD has also been demonstrated to be a key mediator of cyst development and proliferation in polycystic kidney disease (PKD). Therefore, the main objective of my proposed postdoctoral research direction is to characterize the role of NBCe1A as a novel regulator of cystogenesis through activation of sAC-cAMP/PKA signaling pathway. Accordingly, Specific Aim 2 (K00) will test the hypothesis that impaired NBCe1A-mediated HCO3- reabsorption activates a soluble adenylyl cyclase-cAMP/PKA signaling cascade in the collecting duct promoting proliferation and cystogenesis in models of PKD. Together, the F99 and K00 will propel me to achieve my long-term goal to lead an independent research program in nephrology.