SELENOPROTEINS, NF-KB, AND HIV DISEASE IN IV DRUG USERS

在1998年NIDA主办的一次讲习班上，确定了进一步研究营养和代谢因素在艾滋病毒/药物滥用中的作用的必要性。审查的证据包括数据显示，静脉注射吸毒者普遍存在营养不良和微量矿物质硒(Se)和其他抗氧化剂水平低的情况，这些吸毒者还因使用静脉注射药物而表现出氧化应激和脂质过氧化增加。由于血清Se在HIV/AIDS中的进行性下降得到了很好的证明，而且Se状态与HIV疾病进展之间的相关性已经确立，注射吸毒者作为一个群体面临着更高的HIV相关发病率和死亡率加速的风险。值得注意的是，一些病毒，包括痘病毒和HIV-1(我们的初步数据)，编码谷胱甘肽过氧化物酶(GPX)的同源物，GPX是典型的哺乳动物硒蛋白，其中硒半胱氨酸由UGA密码子编码。这与HIV-1尤其相关，因为最近的研究表明，两种细胞硒蛋白GPX和硫氧还蛋白还原酶(TDR)是转录因子NF-kappaB的有效调节者，而转录因子NF-kappaB反过来调节HIV-1基因的表达。我们的初步理论和实验数据表明，一个新的HIV-1基因env-fs被编码在env基因的重叠阅读框架中，其编码的蛋白质是一个高度截断的依赖于Se的GPx模块，在体外实验中具有酶活性。我们还在HIV-1的蛋白水解酶和nef编码区确定了其他几个活性移码位点和潜在的UGA抑制位点；后者与TDR的氧化还原中心有明显的序列相似性。这项研究将评估这些病毒硒蛋白与Se状态和HIV-1疾病进展之间的联系的假设。利用体外方法，具体目标是A1：证明HIV-1基因相关区域编码具有明显生物活性的功能硒蛋白，并且这些新的蛋白质或异构体(例如扩展的nef TDR同源物)实际上在感染细胞中表达；A2：研究预测的病毒硒蛋白(GPX、扩展的nef、Pro-f)对艾滋病毒-1基因表达和氧化应激激活的影响及相互作用；A3：证明病毒硒蛋白合成导致先前在HIV-1感染细胞中证明的细胞硒蛋白水平下降，并且感染细胞的补硒可以部分逆转这一影响。长期目标是为在艾滋病毒吸毒者中使用硒补充剂作为化学保护剂建立分子基础。

At a 1998 NIDA-sponsored workshop, the need for further research on the role of nutritional and metabolic factors in HIV/drug abuse was established. The evidence reviewed included data showing that malnutrition and low levels of the trace mineral selenium (Se) and other antioxidants are commonly evident in IV drug users (IDUs), who also show increased oxidative stress and lipid peroxidation as a consequence of IV drug use. Because a progressive decline in serum Se is well documented in HIV/AIDS, and correlations between Se status and HIV disease progression are firmly established, IDUs as a group are at increased risk for accelerated HIV-related morbidity and mortality. Significantly, some viruses, including a pox virus and HIV-1 (our preliminary data), encode homologues of glutathione peroxidase (GPx), the prototypical mammalian selenoprotein, in which selenocysteine is encoded by the UGA codon. This is particularly relevant for HIV- 1, because recent studies have shown that two cellular selenoproteins, GPx and thioredoxin reductase (TDR), are potent regulators of transcription factor NF-kappaB, which in turn regulates HIV-1 gene expression. Our preliminary theoretical and experimental data show that a novel HIV-1 gene, env-fs, is encoded in an overlapping reading frame of the env gene, and that the protein it codes for is a highly truncated Se-dependent GPx module, enzymatically active in in vitro assays. We have also identified several other active -1 frameshift sites and potential UGA suppression sites in HIV-1, in the protease and nef coding regions; the latter site has clear sequence similarities to the redox centers of TDR. This study will assess the hypothesis that these viral selenoproteins are involved in the connection between Se status and HIV-1 disease progression. Using in vitro methods, the specific aims are A1: to show that the relevant HIV-1 gene regions encode functional selenoproteins that have demonstrable biological activity, and that these novel proteins or isoforms (e.g. the extended nef TDR homolog) are actually expressed in infected cells; A2: to study the effects and interactions of the predicted viral selenoproteins (GPx, extended nef, pro-fs) on HIV-1 gene expression and activation by oxidative stress; A3: to show that viral selenoprotein synthesis contributes to the decline in cellular selenoprotein levels that has been previously demonstrated in HIV-1 infected cells, and that Se supplementation of infected cells can partially reverse this effect. The long- term goal is to establish a molecular basis for the use of Se supplements as a chemoprotectant in HIV+ drug users.