Laboratory for Biosustainability, Institute of Environmental Biology, Wrocław University of Environmental and Life Sciences, Wrocław, Poland.
Microb Cell Fact. 2023 Aug 16;22(1):155. doi: 10.1186/s12934-023-02161-8.
Yarrowia lipolytica is a dimorphic fungus, which switches from yeast to filament form in response to environmental conditions. For industrial purposes it is important to lock cells in the yeast or filamentous form depending on the fermentation process. yl-Hog1 kinase is a key component of the HOG signaling pathway, responsible for activating the osmotic stress response. Additionally, deletion of yl-Hog1 leads to increased filamentation in Yarrowia lipolytica, but causes significant sensitivity to osmotic stress induced by a high concentration of a carbon source.
In this study, we tested the effect of point mutations on the function of yl-Hog1 protein kinase. The targets of modification were the phosphorylation sites (T171A-Y173A) and the active center (K49R). Introduction of the variant HOG1-49 into the hog1∆ strain partially improved growth under osmotic stress, but did not recover the yeast-like shape of the cells. The HOG1-171/173 variant was not functional, and its introduction further weakened the growth of hog1∆ strains in hyperosmotic conditions. To verify a genetic modification in filament form, we developed a new system based on green fluorescent protein (GFP) for easier screening of proper mutants.
These results provide new insights into the functions of yl-Hog1 protein in dimorphic transition and constitute a good starting point for further genetic modification of Y. lipolytica in filament form.
解脂耶氏酵母是一种二相真菌，它会根据环境条件从酵母形态切换到丝状形态。对于工业用途而言，根据发酵过程将细胞锁定在酵母或丝状形态非常重要。yl-Hog1 激酶是 HOG 信号通路的关键组成部分，负责激活渗透胁迫反应。此外，yl-Hog1 的缺失会导致解脂耶氏酵母丝状化增加，但会导致对高浓度碳源诱导的渗透胁迫的敏感性显著增加。
在这项研究中，我们测试了点突变对 yl-Hog1 蛋白激酶功能的影响。修饰的靶点是磷酸化位点（T171A-Y173A）和活性中心（K49R）。将变体 HOG1-49 引入 hog1∆ 菌株中部分改善了渗透压胁迫下的生长，但没有恢复细胞的酵母样形态。HOG1-171/173 变体没有功能，其引入进一步削弱了 hog1∆ 菌株在高渗条件下的生长。为了验证丝状形态的遗传修饰，我们开发了一个基于绿色荧光蛋白（GFP）的新系统，用于更容易筛选合适的突变体。
这些结果为 yl-Hog1 蛋白在二相转变中的功能提供了新的见解，并为进一步在丝状形态中对解脂耶氏酵母进行遗传修饰奠定了良好的基础。
