Molecular Pathways of Programmed C ell Death And Viral C

内部死亡程序控制着体内细胞的数量和类型。疾病可能是由于细胞死亡效率低下或不适当或过度死亡造成的，例如艾滋病期间的人类免疫缺陷病毒（HIV）或SARS期间的SARS冠状病毒造成的死亡。在这个项目中，我们正在采取多方面的方法来研究淋巴细胞和其他细胞类型的凋亡和非凋亡性死亡程序的分子机制。我们研究的主要焦点是肿瘤坏死因子受体（TNFR）超家族中诱导死亡的细胞表面受体，如TNFR1和CD95/Fas/APO-1。这两种受体在刺激细胞凋亡和非凋亡性死亡中发挥重要作用，主要是在免疫过程中。有趣的是，这两种受体在死亡之外都有作用，比如诱导转录因子。我们正试图了解这些受体是如何刺激细胞内机制导致细胞死亡而不是其他细胞结果的。我们把大部分精力集中在一种叫做caspase-8的蛋白酶的激活上，它调节着死亡程序。我们已经描述了两种来自TNFR1和Fas受体的死亡程序，一种是caspase-8依赖性的，具有凋亡形态，另一种是caspase-8独立的，涉及坏死。有趣的是，后一种死亡程序仅在caspase-8被抑制时观察到。这两种形式的淋巴细胞死亡的调控和分子途径是不同的。此外，我们发现抑制非淋巴样细胞中的caspase-8可导致另一种形式的细胞死亡，表现出特殊的细胞质膜结构，称为自噬。尽管最初存在争议，但一些实验室现在已经表明，这种死亡形式对于化疗药物导致肿瘤细胞死亡尤为重要。我们现在已经证明，自噬死亡程序的机制是过氧化氢酶的选择性降解，导致活性氧的明显过度积累，导致细胞损伤和死亡。

Internal death programs control the number and types of cells in the body. Diseases can result from inefficient cell death or from inappropriate or excessive death such as is caused by the human immunodeficiency virus (HIV) during AIDS or the SAR-CoV virus during SARS. In this project we are taking a multifaceted approach to studying molecular mechanisms of both apoptotic and nonapoptotic death programs in lymphocytes as well as other cell types. A major focus of our investigations are death-inducing cell surface receptors in the tumor necrosis factor receptor (TNFR) superfamily such as TNFR1 and CD95/Fas/APO-1. Both receptors play an important role in stimulating both apoptotic and nonapoptotic death of cells principally in immune processes. interestingly, both receptors can have effects beside death such as the induction of transcription factors. We are trying to understand how these receptors stimulate the intracellular machinery that causes cell death in preference to other cellular outcomes. We have focused much of our efforts on the activation of a protease called caspase-8 which regulates the death program. We have characterized two death programs that emanate from TNFR1 and the Fas receptor, one which is caspase-8 dependent and has an apoptotic morphology and the other which is caspase-8 independent and involves necrosis. Interestingly, the latter death program is only observed when caspase-8 is inhibited. The regulation and molecular pathways of these two forms of lymphocyte death are distinct. In addition, we have discovered that inhibition of caspase-8 in non-lymphoid cells can lead to another form of cell death exhibiting particular cytoplasmic membrane structures called autophagy. although initially controversial, several labs have now shown that this form of death is particularly important for the demise of tumor cells by chemotherapeutic agents. We have now shown that the mechanism of autophagic death program is selective degradation of catalase which leads to a marked overaccumulation of reactive oxygen species leading to cellular damageand death. In parallel, we are exploring how the regulation of cellular death programs may play a role in cytopathicity associated with the virus infections in AIDS and SARS. In particular, a critical effect in the onset of AIDS following infection with HIV is the death of T lymphocytes caused by the virus. We have found that this death process is necrotic rather than apoptotic and have now identified two viral gene products, vif and vpr, that are involved in this process. In order to study this process rigorously we have constructed a mathematical model to analyze cell death in tissue culture during HIV infection. Remarkably, both of these cytotoxic gene products cause says cycle arrest at the boundary of the G2 and M phases. The mathematical model reveals that the principal cause of cell loss is cell death rather than cell cycle arrest. We are using molecular genetic approaches to determine if cell cycle arrest actually causes cell death and how this might come about. In contrast to HIV, the human coronavirus that causes SARS, SARS-CoV, causes necrotic cell death that does not involve cell cycle arrest. We have found that cell death can be traced to a novel open reading frame, termed ORF 3b, that is present in SARS-CoV but not other less pathological human coronaviruses. The cellular effect of ORF3b is to cause a dramatic reorganization of the Golgi apparatus which has lethal effects on the cell. We are now trying to established a molecular pathway entrained to ORF3b that causes this cytopathic effect.