巴斯德毕赤酵母SUT2和ATG26响应甲醇代谢机制

Mechanism of SUT2 and ATG26 in response to methanol metabolism in Komagataella phaffii

为寻找巴斯德毕赤酵母(Komagataella phaffii)中甲醇代谢潜在调控因子,通过转录组数据分析,发现受甲醇显著诱导而被甘油所抑制的SUT2(Sterol up take 2,固醇摄取基因2)。固醇是过氧化物酶体膜重要组成部分,过氧化物酶体是甲醇代谢重要场所。为探究SUT2参与甲醇代谢功能机制,通过敲除及过表达SUT2基因,检测K.phaffii在不同碳源、时间点、氧含量条件下胞内麦角固醇含量及AOX1(Alcohol oxidase 1)酶活性变化;发现在甲醇培养基中SUT2通过促进胞内麦角固醇合成及过氧化物酶体膜形成,参与甲醇代谢。与之相反,ATG26(Autophagy-related gene 26,自噬相关基因26)通过促进麦角固醇合成甾醇葡萄糖苷,减少过氧化物酶体膜形成。最后,建立K.phaffii中SUT2及ATG26调控过氧化物酶体膜麦角固醇含量,响应甲醇代谢初步模型。

SUT2(Sterol up take 2)was found as a potential regulator of methanol metabolism in K.phaffii through transcriptome data analysis.SUT2 was significantly induced by methanol and depressed by glycerol.Sterol is an important part of peroxisome membrane,and the peroxisome is an important place for methanol metabolism.The experiment aimed to explore the function of SUT2 in methanol metabolism.SUT2 was knocked out and overexpressed.The intracellular ergosterol content and AOX1(Alcohol oxidase 1)enzyme activity were detected under different carbon sources,different time points and different oxygen contents.It was found that SUT2 was involved in methanol metabolism through promoting the synthesis of intracellular ergosterol and peroxisome membrane.In contrast.ATG26(Autophagy related 26)reduced the formation of peroxisome membrane by promoting the synthesis of sterol glucoside from ergosterol.Finally,a preliminary model was built:SUT2 and ATG26 responsed the methanol metabolism through regulating the ergosterol content of peroxisome membrane in K.phaffii.