LIFE CYCLE REGULATION OF PNEUMOCYSTIS CARINII BY MAPK

MAPK 对卡氏肺囊虫生命周期的调控

• 批准号: 2886123
• 负责人: CHARLES FRANCIS THOMAS
• 金额: $ 7.56万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-01 至 2001-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2886123
• 关键词: Pneumocystis carinii; biological signal transduction; cell cycle; cell differentiation; cell growth regulation; cell proliferation; enzyme activity; flow cytometry; fungal genetics; immunoelectron microscopy; immunoprecipitation; laboratory rat; life cycle; mitogen activated protein kinase; northern blottings; polymerase chain reaction; transfection

Pneumocystis carinii is an opportunistic fungal pathogen which causes severe pneumonia in patients with an impaired immune system, particularly in patients with AIDS. The life cycle of P. carinii is poorly understood. In closely related fungi, Cdc2 associated with a regulatory cyclin controls the transition through the G1 restriction point START in the cell cycle.1-7 In fungi homologous to P. carinii, pheromone mating factors inhibit START through a mitogen-activated protein kinase (MAPK) signal transduction cascade which results in cell cycle arrest and cellular differentiation.8-14 MAPK phosphorylates an inhibitor of the Cdc2-cyclin complex.15-18 MAPK is the essential molecule in the pheromone mating cascade which controls the transition from mitotic growth to a pathway of conjugation, cellular differentiation, and proliferation.8 As an essential molecule regulating an organism's life cycle, MAPK is regulated by a kinase cascade and is activated by phosphorylation on two conserved threonine and tyrosine amino acids.19-24 Attachment of P. carinii to the alveolar epithelium is a central feature of P. carinii pneumonia, and binding to the epithelium is required for the organisms to proliferate.25-26 Our initial studies indicate that P. carinii possesses a MAPK which is highly homologous to fungal MAPK's involved in mating.27 We, therefore, hypothesize that the MAPK signal transduction cascade has a central role in the regulation of the P. carinii life cycle. We further propose that the P. carinii MAPK regulates cellular differentiation and proliferation of P. carinii organisms. To evaluate these hypotheses, we will undertake several parallel investigations. We will characterize the P. carinii MAPK genes involved in life cycle regulation. We will determine the kinase activity and the regulatory phosphorylations for P. carinii MAPK. Further, we will determine whether the expression and activity of P. carinii MAPK is differentially regulated throughout the life cycle of P. carinii by evaluating MAPK in separated cysts and trophozoites. Finally, we will determine the role of MAPK in the transition from one life form to another by evaluating the expression and activity of P. carinii MAPK before and after attachment to the alveolar epithelium and by overexpressing MAPK genes in separated life cycle forms of P. carinii. Through these investigations, we hope to gain important insights into the life cycle regulation of P. carinii.

卡氏肺囊虫是一种机会性真菌病原体，可引起 免疫系统受损的严重肺炎患者， 尤其是艾滋病患者。 P. carinii 的生命周期是 不太了解。 在密切相关的真菌中，Cdc2 与 调节细胞周期蛋白通过 G1 限制控制转变 细胞周期中的起始点1-7 在与 P. carinii 同源的真菌中， 信息素交配因子通过丝裂原激活抑制 START 蛋白激酶 (MAPK) 信号转导级联，导致细胞 周期停滞和细胞分化。8-14 MAPK 磷酸化 Cdc2-cyclin 复合物的抑制剂。15-18 MAPK 是必不可少的 信息素交配级联中控制转变的分子 从有丝分裂生长到接合途径，细胞 分化和增殖。8 作为重要分子 MAPK 调节生物体的生命周期，由激酶调节 级联并通过两个保守苏氨酸的磷酸化被激活 和酪氨酸氨基酸。19-24 P. carinii 与肺泡的附着 上皮细胞是卡氏疟原虫肺炎的一个核心特征，并且与 上皮细胞是生物体增殖所必需的。25-26 初步研究表明 P. carinii 拥有 MAPK 与参与交配的真菌 MAPK 高度同源。27 因此，我们， 假设 MAPK 信号转导级联具有核心作用 参与 P. carinii 生命周期的调节。 我们进一步建议 P. carinii MAPK 调节细胞分化和增殖 P. carinii 生物体。 为了评估这些假设，我们将 进行多项平行调查。我们将描述 P 的特征。 carinii MAPK 基因参与生命周期调控。 我们将确定 P. carinii 的激酶活性和调节性磷酸化 映射。 此外，我们将确定表达和活性是否 P. carinii MAPK 在整个生命周期中受到差异性调节 通过评估分离的包囊和滋养体中的 MAPK 来鉴定 P. carinii。 最后，我们将确定 MAPK 在从一种转变中的作用 通过评估 P 的表达和活性来将生命形式转变为另一种生命形式。 carinii MAPK 附着于肺泡上皮之前和之后 通过在 P. 的不同生命周期形式中过度表达 MAPK 基因。 卡里尼。 通过这些调查，我们希望获得重要的 对 P. carinii 生命周期调控的见解。