Complementation and characterization of the Pneumocystis carinii MAPK, PCM.
Complementation and characterization of the Pneumocystis carinii MAPK, PCM.
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卡氏肺囊虫 MAPK、PCM 的补充和表征。
DOI：
10.1016/s0014-5793(03)00914-1
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发表时间：
2003
期刊：
影响因子：
3.5
通讯作者：
ThomasJr,CharlesF
中科院分区：
文献类型：
--
作者：
Vohra,PawanK;Puri,Veenu;ThomasJr,CharlesF
丝裂原活化蛋白激酶(MAPK)途径将环境信号传递到细胞内，如增殖和分化。真菌利用特定的信息素诱导的MAPK途径来调节接合、子囊的形成和进入减数分裂。我们之前已经从真菌机会主义的肺孢子虫中发现了一种MAPK，PCM，它导致了艾滋病患者的严重肺炎。为了深入了解PCM的功能，我们在信息素信号缺乏的酿酒酵母中表达了PCM，并检测了该MAPK通路的激活和抑制情况。PCM可使酿酒酵母Fus3ΔKss1Δ突变株的信息素信号得到恢复(提高6倍)，且不受渗透胁迫的影响。通过该途径的信号在10μ的MU0126中减少2.5倍，但不受SB203580的影响。我们对从卡氏肺孢子虫中分离的天然PCM激酶的活性条件进行了评估，发现最适的条件是0.1 mM的氯化镁，pH 6.5，温度30-35℃，10μM的三磷酸腺苷。与包囊相比，营养型卡氏肺孢子虫的PCM活性显著升高，这表明PCM在卡氏肺孢子虫从营养型向包囊的生命周期转变中发挥了作用。
Mitogen-activated protein kinase (MAPK) pathways transfer environmental signals into intracellular events such as proliferation and differentiation. Fungi utilize a specific pheromone-induced MAPK pathway to regulate conjugation, formation of an ascus, and entry into meiosis. We have previously identified a MAPK, PCM, from the fungal opportunist Pneumocystis, responsible for causing severe pneumonia in patients with AIDS. In order to gain insight into the function of PCM, we expressed it in Saccharomyces cerevisiae deficient in pheromone signaling and tested activation and inhibition of this MAPK pathway. PCM restored pheromone signaling in S. cerevisiae fus3Δ kss1Δ mutants with α-factor pheromone (six-fold increase) and was not activated by osmotic stress. Signaling through this pathway decreased 2.5-fold with 10 μM U0126, and was unaffected with SB203580. We evaluated the conditions for native PCM kinase activity isolated from Pneumocystis carinii organisms and found that 0.1 mM MgCl2, pH 6.5, temperature 30–35°C, and 10 μM ATP were optimal. The activity of PCM is significantly elevated in P. carinii trophic forms compared to cysts, implicating a role for PCM in the life cycle transition of P. carinii from trophic forms to cysts.