Regulation of heme oxygenase-1 by biliverdin reductase

说明(申请人提供)：变性的血红素蛋白(血红素蛋白)的血红素部分被HO-1和HO-2降解为CO和HO活性抑制剂胆绿素(BV)；BV被其还原酶(BVR)还原为抗氧化剂胆红素。CO具有抗炎和血管扩张活性。引起氧化应激和血球蛋白变性的刺激因素，如外科手术、遗传性和传播性溶血性疾病，以及某些药物、工业和环境制剂，都会导致HO-1。肾小管是血红素化合物强大的促氧化活性的靶标。到目前为止，还没有有效的策略来对抗血红素蛋白的肾毒性；然而，通过激活应激激活的反应元件，如AP-1/CRE，ARES(抗氧化反应元件)，增加HO-1的活性被认为是细胞保护。这种激活涉及碱性亮氨酸拉链(BZip)转录因子c-jun、ATF-2/CREB和Nrf2的细胞独立结合。MAPK和PI3-K通路利用PKCs传递激活bZip因子和“串扰”的信号。BZip因子的活性取决于其二聚体伙伴的身份。最终靶基因产物如HO-1的磷酸化改变其活性和周转。在体外和培养的细胞中，我们发现：人(H)BVR是控制MAPK和PI3-K信号转导的罕见激酶之一；是一个bZip因子；被HO-1诱导剂激活；以及胞浆和细胞核之间的交通。HBVR与AP-L/Cre和ARE元件结合，也增强ATF-2和NRF2与AP-1和/或ARE的结合；促进ATF-2、c-jun和c-Fos的诱导和激活；激活HO-1反应的激酶介质，即PKCs和PKB/Akt；并导致细胞分化。BVR调节HO-1氧化应激反应及其抗细胞凋亡作用。这项应用的总体目标是在分子和细胞水平上进一步研究BVR对HO-1活性的调节，并将研究扩展到完整的动物。具体目的是检测：1)BVR作为激酶的功能：在HO-1的磷酸化、活性和周转中的激酶；2)hBVR在ATF-2和Nrf2转录活性中的作用；以及，3)增加BVR水平是否对血红素介导的损伤具有保护作用。在肾小管中表达hBVR并用溶血剂苯肼治疗的小鼠将被分析病理生理学、抗氧化状态、激酶活性和HO-1水平。肝脏将作为控制转基因表达的非靶器官。

DESCRIPTION (provided by applicant): The heme moiety of denatured hemoproteins (heme-proteins) is degraded by HO-1 & HO-2, to CO and biliverdin (BV), an HO activity inhibitor; BV is reduced by its reductase (BVR) to the antioxidant, bilirubin. CO has anti-inflammatory and vasodilatory activities. Stimuli that cause oxidative stress and hemoprotein denaturation, such as surgical interventions, inherited and transmitted hemolytic diseases, and certain drugs, industrial and environmental agents, induce ho-1. The kidney tubules are the target of potent pro-oxidant activity of heme-compounds. To date, no effective strategy has been described to counter heme-protein renal toxicity; however, increase in HO-1 activity by activation of stress-activated response elements, e.g., AP-1/CRE, AREs (antioxidant response elements) is considered cytoprotective. The activation involves cell-line independent binding of basic leucine zipper (bZip) transcription factors: c-Jun, ATF- 2/CREB, and Nrf2. The MAPK and PI3-K pathways transduce signals for activation of bZip factors and "cross talk" using PKCs. bZip factor activity is subject to the identity of its dimeric partner. Phosphorylation of the ultimate target gene product, e.g., HO-1, alters its activity and turnover. In vitro and in cultured cells, we have discovered that: the human (h) BVR is one of the rare kinases that control MAPK and PI3-K signaling; is a bZip factor; activated by ho-1 inducers; and, traffics between the cytosol and nucleus. hBVR binds to AP-l/CRE and ARE elements and also enhances ATF-2 and Nrf2 binding to AP-1 and/or ARE; promotes induction and activation of ATF-2, c-Jun and c-Fos; activates kinase mediators of ho-1 response, i.e. PKCs and PKB/Akt; and, causes cell differentiation. BVR regulates ho-1 oxidative stress response and its anti-apoptotic effect. The overall objective of this application is to further investigate regulation of HO-1 activity by BVR at the molecular and cellular levels and to extend the investigation to the intact animal. Specific aims are to examine: i) function of BVR as kinase:kinase in phosphorylation, activity, and turnover of HO-1; 2) the role of hBVR in transcriptional activity of ATF-2 and Nrf2; and, 3) whether increased levels of BVR protect against heme-mediated injury. Mice expressing hBVR in renal tubules and treated with the hemolytic agent, phenylhydrazine, will be analyzed for pathophysiology, antioxidant status, kinase activities and HO-1 levels. Liver will serve as the non-target organ as control for the expression of the transgene.