Stress sensitivity of a fission yeast strain lacking histidine kinases is rescued by the ectopic expression of Chk1 from Candida albicans

Current Genetics, Sep 2016

The development of new drugs against the pathogenic yeast Candida albicans is compelling and the evolution of relevant bioassays is important to achieve this goal. Promising drug targets are proteins that lack human counterparts which are true for the His-to-Asp phosphorelay signal transduction systems, important for stress sensing in bacteria, fungi, and plants. In the pathogenic yeast, Candida albicans, the CaChk1 histidine kinase is a trigger of the pathway that leads to a switch from yeast to hyphal growth necessary for invasion. Intriguingly, the model yeast Schizosaccharomyces pombe has a similar phosphorelay system, with three histidine kinases named Mak1, Mak2, and Mak3, which are important for the prevention of aberrant mating and sporulation on rich media. This study uncovered distinct functions for the three histidine kinases; Mak1 alone or Mak2 and Mak3 together were sufficient for the repression of the meiotic cycle when nutrients were available. Moreover, strains lacking histidine kinase genes were sensitive to various types of stress conditions in an auxotrophic strain background, while the stress sensitivity was lost in prototrophic strains. Finally, the stress sensitivity of a S. pombe strain that lacks endogenous histidine kinases could be complemented by the ectopic expression of the CaChk1 histidine kinase from C. albicans. This finding opens up for the possibility to perform a drug screen with a biological read-out in S. pombe to find inhibitors of CaChk1.

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Stress sensitivity of a fission yeast strain lacking histidine kinases is rescued by the ectopic expression of Chk1 from Candida albicans

Stress sensitivity of a fission yeast strain lacking histidine kinases is rescued by the ectopic expression of Chk1 from Candida albicans Vladimir Maksimov 0 1 2 Marcus Wäneskog 0 1 2 Alejandro Rodriguez 0 1 2 Pernilla Bjerling 0 1 2 0 Present Address: Dept. of Cell and Molecular Biology, University of Uppsala , Box 596, 751 24 Uppsala , Sweden 1 Science for Life Laboratory, Department of Medical Biochemistry and Microbiology (IMBIM), University of Uppsala , Box 582, 751 23 Uppsala , Sweden 2 Communicated by M. Kupiec 3 Pernilla Bjerling The development of new drugs against the pathogenic yeast Candida albicans is compelling and the evolution of relevant bioassays is important to achieve this goal. Promising drug targets are proteins that lack human counterparts which are true for the His-to-Asp phosphorelay signal transduction systems, important for stress sensing in bacteria, fungi, and plants. In the pathogenic yeast, Candida albicans, the CaChk1 histidine kinase is a trigger of the pathway that leads to a switch from yeast to hyphal growth necessary for invasion. Intriguingly, the model yeast Schizosaccharomyces pombe has a similar phosphorelay system, with three histidine kinases named Mak1, Mak2, and Mak3, which are important for the prevention of aberrant mating and sporulation on rich media. This study uncovered distinct functions for the three histidine kinases; Mak1 alone or Mak2 and Mak3 together were sufficient for the repression of the meiotic cycle when nutrients were available. Moreover, strains lacking histidine kinase genes were sensitive to various types of stress conditions in an auxotrophic strain background, while the stress sensitivity was lost in prototrophic strains. Finally, the stress sensitivity of a S. pombe strain that lacks endogenous histidine kinases could be complemented by the ectopic expression of the CaChk1 histidine kinase from C. albicans. This finding opens up for the possibility to perform a drug screen with a biological read-out in S. pombe to find inhibitors of CaChk1. Histidine kinase; Candida albicans; Fission yeast; Schizosaccharomyces pombe; His-to-Asp phosphorelay; Stress response - V. Maksimov and M. Wäneskog contributed equally. Introduction Proteins in pathogenic microorganisms that lack human counterparts are highly interesting for the development of new drugs. In bacteria, fungi, and plants, but not in animals, histidine kinases (HK) are part of a His-to-Asp phosphorelay signal transduction system. In bacteria, it is a two-component system, where the HK phosphorylates an aspartic acid residue on a receiver protein. In fungi, the system typically involves three components: a HK, an intermediate Histidine Phosphotransfer (HPt) protein, and a downstream Response Regulator (RR) protein (Casino et al. 2010) . Quite a lot of effort has been done to find inhibitors of bacterial HKs to develop novel antibiotics, but only limited screens has been made with fungal HKs as target (Tebbets et al. 2012, 2013; Bem et al. 2015) . The fungal signal system has mostly been studied in budding yeast, Saccharomyces cerevisiae, where there is one HK (ScSln1), one HPt (ScYpd1), and two RR (ScSsk1 and ScSkn7) (Brown et al. 1994; Posas et al. 1996; Li et al. 1998) . In budding yeast, phosphotransfer occurs constitutively during normal growth conditions, which results in a phosphorylated ScSsk1 protein that represses the downstream Hog1 Mitogen Activated Protein Kinases (MAPK) pathway. In contrast, during osmotic stress, the unphosphorylated ScSsk1 protein somehow activates the MAPK pathway resulting in transcriptional regulation that leads to adaptation to high osmolarity (Posas et al. 1996) . Deletion of SLN1 or YPD1 in budding yeast is lethal, since it results in a constitutive activation of the Hog1 MAPK pathway. In fission yeast, Schizosaccharomyces pombe, there is a similar phosphorelay system with some variations from budding yeast. First, there are three HKs, Mak1 (Phk3), Mak2 (Phk1), and Mak3 (Phk2); second, knocking out all three HKs is not lethal in fission yeast. Mak2 and Mak3 have been reported to be involved in signal transduction during oxidative stress, although a mak2,3Δ double mutant strain is not sensitive to hydrogen peroxide (Buck et al. 2001) . In fission yeast, there is one HPt protein, Mrp1, that functions together with the HKs in response to free radicals (Nguyen et al. 2000; Buck et al. 2001; Tan et al. 2007) . The HPt protein transfers the phosphate to one of the two RR proteins named Mcs4 and Prr1 (Buck et al. 2001; Quinn et al. 2011; Morigasaki and Shiozaki 2013) . The details of the phosphotransfer in the two-component system and the subsequent activation of the MAPK pathway have not been investigated in S. pombe, but it is documented that when cells are grown vegetatively on rich media, the MAPK signalling pathway is repressed and the HKs are required for that repression (Nakamichi et al. 2002) . When nitrogen is depleted from (...truncated)


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Vladimir Maksimov, Marcus Wäneskog, Alejandro Rodriguez, Pernilla Bjerling. Stress sensitivity of a fission yeast strain lacking histidine kinases is rescued by the ectopic expression of Chk1 from Candida albicans, Current Genetics, 2017, pp. 343-357, Volume 63, Issue 2, DOI: 10.1007/s00294-016-0644-9