【摘要】 红曲菌(Monascus)是我国传统的发酵菌株，在发酵过程中会产生一种具有肾毒性的真菌毒素——桔霉素(Citrinin, CIT)。桔霉素的存在极大地限制了红曲制品的应用与发展。明确红曲菌合成CIT的机制，对于开发抑制CIT合成的方法具有十分重要的意义。小G蛋白Rab1可以介导调控丝状真菌中蛋白的转运，ctnE基因编码红曲菌AS3.4384中CIT合成基因簇中的脱氢酶。然而，目前尚不清楚Rab1蛋白是否参与了CtnE蛋白的转运。通过农杆菌介导转化的方法构建了AS3.4384::Rab1-GFP(绿色荧光蛋白，Green Fluorescent Protein)和AS3.4384::Rab1-GFP::ctnE-mCherry(红色荧光蛋白，monomeric cherry red fluorescent protein)两种红曲菌株，并通过荧光共定位确定了Rab1蛋白和CtnE蛋白在菌体中的分布情况。结果表明，CtnE-mCherry蛋白大多聚集于红曲菌丝体的顶端以及分生孢子中，并在菌丝体内呈均匀弥散状分布。Rab1蛋白和CtnE蛋白在红曲菌体内的分布位点基本一致，表明CIT的合成与囊泡转运密切相关。通过Rab1蛋白与CtnE蛋白的荧光共定位情况进一步解析了CIT合成位点。更多还原

【Abstract】 Monascus,a traditional fermentation strain in China, produces a nephrotoxic fungal toxin called citrinin(CIT) during fermentation.The presence of citrinin greatly limits the application and development of Monascus products.Understanding the mechanism of CIT synthesis by Monascus is of great significance for developing methods to inhibit its synthesis.The small G protein Rab1 can mediate the regulation of protein transport in filamentous fungi, and the ctnE gene encodes a dehydrogenase in the CIT biosynthetic gene cluster of Monascus AS3.4384.However, it is currently unclear whether the Rab1 protein is involved in the transport of the CtnE protein.Two Monascus strains, AS3.4384::Rab1-GFP(Green Fluorescent Protein) and AS3.4384::Rab1-GFP::ctnE-mCherry(monomeric cherry red fluorescent protein),were constructed using Agrobacterium-mediated transformation method.Fluorescence co-localization was used to determine the distribution of Rab1 and CtnE proteins in the fungal cells.The results showed that the CtnE-mCherry protein mostly accumulated at the tips of Monascus hyphae and in the developing spores, with a uniform distribution within the hyphae.The distribution sites of Rab1 and CtnE proteins in Monascus cells were essentially consistent, indicatied a close relationship between CIT synthesis and vesicle transport.Further analysis of the fluorescence co-localization of Rab1 and CtnE proteins elucidated the sites of CIT synthesis.更多还原