Ting S Y, Ishola O A, Ahmed M A, Tabana Y M, Dahham S, Agha M T, Musa S F, Muhammed R, Than L T L, Sandai D
Infectomics Cluster, advanced medical and dental institute, universiti Sains Malaysia, Jln Tun Hamdan Sheikh Tahir, 13200 Bertam Penang, Malaysia.
Regenerative medicine cluster, advanced medical and dental institute, UniversitiSains Malaysia, 13200 Bertam Penang, Malaysia.
J Mycol Med. 2017 Mar;27(1):98-108. doi: 10.1016/j.mycmed.2016.12.002. Epub 2016 Dec 29.
The virulence of Candida albicans is dependent upon fitness attributes as well as virulence factors. These attributes include robust stress responses and metabolic flexibility. The assimilation of carbon sources is important for growth and essential for the establishment of infections by C. albicans. Previous studies showed that the C. albicans ICL1 genes, which encode the glyoxylate cycle enzymes isocitratelyase are required for growth on non-fermentable carbon sources such as lactate and oleic acid and were repressed by 2% glucose. In contrast to S. cerevsiae, the enzyme CaIcl1 was not destabilised by glucose, resulting with its metabolite remaining at high levels. Further glucose addition has caused CaIcl1 to lose its signal and mechanisms that trigger destabilization in response to glucose. Another purpose of this study was to test the stability of the Icl1 enzyme in response to the dietary sugars, fructose, and galactose. In the present study, the ICL1 mRNAs expression was quantified using Quantitative Real Time PCR, whereby the stability of protein was measured and quantified using Western blot and phosphoimager, and the replacing and cloning of ICL1 ORF by gene recombination and ubiquitin binding was conducted via co-immuno-precipitation. Following an analogous experimental approach, the analysis was repeated using S. cerevisiaeas a control. Both galactose and fructose were found to trigger the degradation of the ICL1 transcript in C. albicans. The Icl1 enzyme was stable following galactose addition but was degraded in response to fructose. C. albicans Icl1 (CaIcl1) was also subjected to fructose-accelerated degradation when expressed in S. cerevisiae, indicating that, although it lacks a ubiquitination site, CaIcl1 is sensitive to fructose-accelerated protein degradation. The addition of an ubiquitination site to CaIcl1 resulted in this enzyme becoming sensitive to galactose-accelerated degradation and increases its rate of degradation in the presence of fructose. It can be concluded that ubiquitin-independent pathways of fructose-accelerated enzyme degradation exist in C. albicans.
白色念珠菌的毒力取决于其适应性特征以及毒力因子。这些特征包括强大的应激反应和代谢灵活性。碳源的同化对于白色念珠菌的生长很重要，也是其建立感染所必需的。先前的研究表明，白色念珠菌中编码乙醛酸循环酶异柠檬酸裂合酶的ICL1基因对于在乳酸和油酸等非发酵碳源上生长是必需的，并且受到2%葡萄糖的抑制。与酿酒酵母不同，CaIcl1酶不会因葡萄糖而不稳定，其代谢产物仍保持在高水平。进一步添加葡萄糖会导致CaIcl1失去信号以及响应葡萄糖触发不稳定的机制。本研究的另一个目的是测试Icl1酶对膳食糖、果糖和半乳糖的稳定性。在本研究中，使用定量实时PCR对ICL1 mRNA的表达进行定量，通过蛋白质印迹和磷成像仪测量和定量蛋白质的稳定性，并通过共免疫沉淀进行基因重组和泛素结合对ICL1开放阅读框进行替换和克隆。采用类似的实验方法，以酿酒酵母作为对照重复分析。发现半乳糖和果糖都会触发白色念珠菌中ICL1转录本的降解。添加半乳糖后Icl1酶稳定，但响应果糖会被降解。当在酿酒酵母中表达时，白色念珠菌Icl1（CaIcl1）也会受到果糖加速降解的影响，这表明，尽管它缺乏泛素化位点，但CaIcl1对果糖加速的蛋白质降解敏感。在CaIcl1上添加泛素化位点会导致该酶对半乳糖加速降解敏感，并在果糖存在的情况下增加其降解速率。可以得出结论，白色念珠菌中存在果糖加速酶降解的非泛素依赖性途径。
