Regulation of the ER-stress regulator IRE1α - a new approach to the modulation of ischemia-induced mitochondrial dysfunction and cell death

缺血再灌注损伤(IRI)引起的组织损伤仍然是致残率和死亡率的主要原因。虽然与IRI相关的组织损伤主要发生在再灌注期间，但在缺血期间触发的不同途径启动了与再灌注相关的损伤，包括细胞内钙流量增加和线粒体应激相关的细胞死亡。根据最近发表的数据和初步工作，我们认为内质网(ER)相关的同源二聚体蛋白IRE1α在缺氧期获得瞬时的单体状态，允许与IP3R1的异二聚化。IRE1VDAC1-α异源二聚体在内质网线粒体相关膜(ER-MAM)处诱导IP3R1VDAC1通道的形成，从而促进钙离子从内质网流向线粒体。这种钙内流启动了再灌流期间线粒体功能障碍和促进线粒体细胞死亡的作用。基于进一步的初步工作，我们认为这种启动缺血细胞损伤的新机制可以被抑制，从而防止IRI相关的细胞和组织损伤。在调节IRE1IP3R1VDAC1异源二聚体诱导的α-VDAC1通道形成的可能候选基因中，激活蛋白C(Apc)就是其中之一。APC是一种具有细胞保护作用的凝血酶，已知通过不明确的细胞内机制来改善IRI。为了研究这一新的机制及其调控，我们希望解决以下目标：目的1：研究IRE1α-IP3R1相互作用的功能相关性和分子基础；目的2：确定调控IRI期间ER-MAM中IRE1α新功能的分子通路；目的3：研究APC和IRE1α在IRI中的功能相关性和机制相互作用；我们期望通过回答这些问题，能够设计有针对性的方法来特异性抑制IRE1α-IP3R1的异构化，从而防止线粒体钙超载和线粒体功能障碍，从而促进细胞损伤。因此，我们期待着对缺血引发的细胞和组织损伤的机制以及调控ER-MAM形成的机制有新的见解。

Tissue damage due to ischemia reperfusion injury (IRI) remains a major cause of morbidity and mortality. While the tissue injury associated with IRI occurs primarily during reperfusion, different pathways triggered during ischemia prime the injury associated with reperfusion, including increased intracellular calcium flux and mitochondrial-stress associated cell death. Based on recently published data and preliminary work we propose that the endoplasmic reticulum (ER) associated homodimeric protein IRE1α acquires a transient monomeric state during the hypoxic phase, allowing heterodimerization with IP3R1. The IRE1α – IP3R1 heterodimer induces IP3R1 – VDAC1 channel formation at the ER-MAMs (ER-mitochondria associated membranes), which facilitates calcium flux from the ER to mitochondria. This calcium influx primes mitochondrial dysfunction and mitochondrial cell-death promoting effects during reperfusion. Based on further preliminary work we propose that this novel mechanism, which primes ischemic cells for damage, can be inhibited, thus preventing IRI-associated cell- and tissue damage. Among possible candidates regulating the IRE1α – IP3R1 heterodimer induced IP3R1-VDAC1 channel formation is activated protein C (aPC). aPC is a coagulation protease with cytoprotective effects, known to ameliorate IRI through poorly defined intracellular mechanisms. To investigate this novel mechanism and its regulation we wish to address the following aims: Aim-1: Investigate the functional relevance and molecular basis of the IRE1α–IP3R1 interaction; Aim-2: Identify the molecular pathways that regulate the new function of IRE1α at the ER-MAMs during IRI; Aim-3: Investigate the functional relevance and mechanistic interaction of aPC with IRE1α in IRI; We expect that answering these questions will enable us to design targeted approaches to specifically inhibit IRE1α – IP3R1 heterodimerization and thus preventing the mitochondrial calcium overload and mitochondrial dysfunction, which primes cells for damage. We thus expect new insights into mechanisms of ischemia-primed cell- and tissue-damage as well as into mechanisms regulating ER-MAM formation.