Candida albicans response regulator gene SSK1 regulates a subset of genes whose functions are associated with cell wall biosynthesis and adaptation to oxidative stress
Candida albicans response regulator gene SSK1 regulates a subset of genes whose functions are associated with cell wall biosynthesis and adaptation to oxidative stress
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DOI：
10.1128/ec.2.5.1018-1024.2003
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发表时间：
2003-10-01
期刊：
影响因子：
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通讯作者：
Calderone, R
中科院分区：
文献类型：
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作者：
Chauhan, N;Inglis, D;Calderone, R
白念珠菌Ssk 1 p是Hog 1双组分信号转导系统中的一个假定的反应调节蛋白。在酿酒酵母中，Ssk 1 p的磷酸化状态决定了促进细胞适应渗透胁迫的基因是否被激活。我们以前已经证明，C。白色念珠菌SSK 1不与S. C. cerevisiae的ssk 1突变体，白色念珠菌对山梨醇不敏感。在这项研究中，我们表明，C。白色念珠菌ssk 1突变体对几种氧化剂敏感，包括过氧化氢、叔丁基氢过氧化物、甲萘醌和超氧化钾，当每种氧化剂掺入酵母提取物-蛋白胨-葡萄糖（YPD）琼脂培养基中时。我们使用DNA微阵列来识别受ssk 1突变影响的基因。将在YPD培养基中于30 ℃生长3小时的突变细胞（CSSK 21株）的RNA逆转录，然后与类似制备的野生型细胞（CAF 2）的RNA进行比较。我们观察到来自突变细胞的七个基因一致上调（与CAF 2相比，三倍或更高）。In S.在酿酒酵母中，上调的七个基因中的三个（AHP 1，HSP 12和PYC 2）为细胞提供了响应氧化应激的适应功能;另一个基因（GPH 1）在应激条件下由Hog 1 p调节。另外三个上调的基因编码细胞表面蛋白（FLO 1）、甘露糖基转移酶（MNN 4 -4）和调节C.白色念珠菌在下调的基因中，ALS 1是白色念珠菌中已知的细胞粘附素。通过对HSP 12、AHP 1、CHK 1、PYC 2、GPH 1、ALS 1、MNN 4 -4和FLO 1的逆转录-PCR获得微阵列数据的验证。为了进一步明确Ssk 1 p在C.在白色念珠菌中，我们使用Western印迹分析来测量Hog 1 p在C. ssk 1突变体中的磷酸化。当生长在渗透压或氧化应激下时，白色念珠菌。我们观察到Hog 1 p在C. ssk 1突变体中被磷酸化。当在高渗培养基中生长时，在白色念珠菌中没有磷酸化，但当在过氧化氢存在下生长时，在SSK 1突变体中没有磷酸化。这些数据表明C.白念珠菌利用Ssk 1 p反应调节蛋白使细胞适应氧化应激，而其在适应渗透应激中的作用不太确定。此外，SSK 1似乎在细胞壁生物合成的某些方面具有调节功能。因此，C. albicans SSK 1与S.酿酒酵母SSK 1.
Ssk1p of Candida albicans is a putative response regulator protein of the Hog1 two-component signal transduction system. In Saccharomyces cerevisiae, the phosphorylation state of Ssk1p determines whether genes that promote the adaptation of cells to osmotic stress are activated. We have previously shown that C. albicans SSK1 does not complement the ssk1 mutant of S. cerevisiae and that the ssk1 mutant of C. albicans is not sensitive to sorbitol. In this study, we show that the C. albicans ssk1 mutant is sensitive to several oxidants, including hydrogen peroxide, t-butyl hydroperoxide, menadione, and potassium superoxide when each is incorporated in yeast extract-peptone-dextrose (YPD) agar medium. We used DNA microarrays to identify genes whose regulation is affected by the ssk1 mutation. RNA from mutant cells (strain CSSK21) grown in YPD medium for 3 h at 30degreesC was reverse transcribed and then compared with similarly prepared RNA from wild-type cells (CAF2). We observed seven genes from mutant cells that were consistently up regulated (three-fold or greater compared to CAF2). In S. cerevisiae, three (AHP1, HSP12, and PYC2) of the seven genes that were up regulated provide cells with an adaptation function in response to oxidative stress; another gene (GPH1) is regulated under stress conditions by Hog1p. Three other genes that are up regulated encode a cell surface protein (FLO1), a mannosyl transferase (MNN4-4), and a putative two-component histidine kinase (CHK1) that regulates cell wall biosynthesis in C. albicans. Of the down-regulated genes, ALS1 is a known cell adhesin in C albicans. Verification of the microarray data was obtained by reverse transcription-PCR for HSP12, AHP1, CHK1, PYC2, GPH1, ALS1, MNN4-4, and FLO1. To further determine the function of Ssk1p in the Hog1p signal transduction pathway in C. albicans, we used Western blot analysis to measure phosphorylation of Hog1p in the ssk1 mutant of C. albicans when grown under either osmotic or oxidative stress. We observed that Hog1p was phosphorylated in the ssk1 mutant of C. albicans when grown in a hyperosmotic medium but was not phosphorylated in the ssk1 mutant when the latter was grown in the presence of hydrogen peroxide. These data indicate that C. albicans utilizes the Ssk1p response regulator protein to adapt cells to oxidative stress, while its role in the adaptation to osmotic stress is less certain. Further, SSK1 appears to have a regulatory function in some aspects of cell wall biosynthesis. Thus, the functions of C. albicans SSK1 differ from those of S. cerevisiae SSK1.