CALCIUM AND PH HOMEOSTASIS IN PNEUMOCYSTIS CARINII

自1981年发现艾滋病以来，卡氏肺孢子虫的发病率 肺炎急剧增加。本文还讨论了对P. 艾滋病中的卡氏感染（例如，高复发率和不良反应率） 药物反应）强调了开发新形式的 疗法我们的研究兴趣涉及一种理性的方法， 化学治疗控制传染源的目的是发现新的 药物作用的目标。我们发现，与 在哺乳动物细胞中，ATP依赖性H+挤出机制是主要的 卡氏疟原虫滋养体用于维持其 细胞内pH（pHi）稳态。我们的假设是， 由H+-ATP酶控制的稳态机制（pH稳态， 营养吸收、Na+和Ca 2+排出）将中断卡氏原螯虾的生命 周期因此，它们的识别可能有助于 开发新的化学治疗剂或化学预防剂， 这些病原体。这项建议的具体目标是：（1） 描述了卡氏肺孢子虫的ATP依赖性H+-挤出机制。 这些研究将涉及对以下物质的Cl-要求的调查： H+挤出; H+-的生电性质的研究 葡萄糖对质膜质子影响的研究 研究质子泵在养分运输中的作用; 确认ATP酶基因的产物， 从卡氏肺孢子虫中克隆并测序的对应于质膜 H+-ATP酶活性，我们已经确定;溶解和 酶的纯化和药物作用的研究 影响卡氏肺孢子虫H+-ATP酶活力的因素;（2） 探讨卡氏肺吸虫PHI与Ca ~（2+）稳态的关系。 我们将研究卡氏肺孢子虫H+-ATP酶在控制 在这些微生物中，如果pHi的变化可以起作用， 触发钙信号，如果pHi和细胞内 Ca ~（2+）浓度可能参与了 介导卡氏疟原虫滋养体与宿主细胞的附着。

Since the discovery of AIDS in 1981, the incidence of P. carinii pneumonia has increased dramatically. The problems of treatment of P. carinii infection in AIDS (e.g. the high rates of relapse and adverse drug reactions) have emphasized the need to develop new forms of therapy. Our research interest involves a rational approach to chemotherapeutic control of infectious agents aimed at finding new targets for drug action. We have identified that, in contrast to mammalian cells, an ATP-dependent H+-extrusion mechanism is the main mechanism that P. carinii trophozoites use for the maintenance of their intracellular pH (pHi) homeostasis. Our hypothesis is that disruption of the homeostatic mechanisms controlled by the H+-ATPase (pH homeostasis, nutrient uptake, Na+ and ca2+ extrusion) would interrupt P. carinii life cycle. Therefore, their identification could be useful for the development of new chemotherapeutic or chemoprophylactic agents against these pathogens. The specific aims of this proposal are: (1) To characterize the ATP-dependent H+-extrusion mechanism of P. carinii. These studies will involve the investigation of the Cl- requirement for H+ extrusion; the investigation of the electrogenic nature of the H+- ATPase; the study of the effect of glucose on the plasma membrane proton pump; the study of the role of the proton pump in nutrient transport; the confirmation that the product of the ATPase gene that has been cloned and sequenced from P. carinii corresponds to the plasma membrane H+-ATPase activity that we have identified; the solubilization and purification of the enzyme; and the investigation of the effect of drugs that affect the P. carinii H+-ATPase on their viability; and (2) To investigate the relation between PHI and ca2+ homeostasis in P. carinii. We will investigate the role of the P. carinii H+-ATPase in the control of Ca2+ homeostasis in these microorganisms, if changes in pHi can serve to trigger calcium signals, and if changes in pHi and in intracellular ca2+ concentration could be involved in the signaling mechanisms that mediate the attachment of P. carinii trophozoites to the host cells.