新生隐球菌SER5基因的敲除与功能验证研究

中国真菌学杂志　 2021, Vol. 16 　Issue (5): 335-340.

新生隐球菌SER5基因的敲除与功能验证研究

童双礼^1,3, 吴洁², 李莎莎⁴, 桑军军^1,4

1. 福建医科大学福总临床医学院(第九○○医院)烧伤整形科, 福州 350001;
2. 福建医科大学福总临床医学院皮肤科, 福州 350001;
3. 32231部队卫生所, 福州 350001;
4. 厦门大学附属东方医院, 福州 350001

收稿日期:2021-02-02 发布日期:2021-10-28
通讯作者: 桑军军,E-mail:sangjunjunsang@163.com E-mail:sangjunjunsang@163.com
作者简介:童双礼,男(汉族),本科,主治医师.E-mail:1186007195@qq.com
基金资助:
福建医科大学启航基金（2017XQ1200）；福建省自然科学基因（2018J05061）

Deletion and functional studies of SER5 gene in Cryptococcus neoformans

Tong Shuangli^1,3, WU Jie², LI Shasha⁴, SANG Junjun^1,4

1. Department of Burn and Plastic Surgery, 900 Hospital, Fuzhou General Clinical Medical College, Fujian Medical University, Fuzhou 350001, China;
2. Department of dermatology, 900 Hospital, Fuzhou General Clinical Medical College, Fujian Medical University, Fuzhou 350001, China;
3. Health-center of 32231 Army, Fuzhou 350001, China;
4. Dongfang Hospital, Xiamen University, Fuzhou 350001, China

Received:2021-02-02 Published:2021-10-28

摘要/Abstract

摘要： 目的构建新生隐球菌SER5基因缺陷株，并初步探究其功能。方法敲除SER5基因，通过体外应激应答、黑色素诱导、荚膜诱导、尿素酶测定、生长曲线测定、酵母双杂交实验考察SER5在新生隐球菌中的功能。结果成功构建SER5基因缺陷株，SER5基因缺失后对新生隐球菌的体外应激、荚膜、分解尿素及生长没有显著的影响，但新生隐球菌黑色素合成有显著的降低，通过酵母双杂交实验筛选出可能与Ser5相互作用的19种不同的蛋白编码基因。结论新生隐球菌Ser5影响新生隐球菌黑色素生成，可能与内质网蛋白、内质网稳定蛋白、VAMP7、磷脂转运蛋白、Gmt1、Vti1a等存在相关作用，提示Ser5参与新生隐球菌黑色素的合成或转运过程。

关键词: 新生隐球菌, 丝氨酸蛋白酶, 黑色素, 功能

Abstract: Objective To construct SER5 gene deletion and reconstruction mutants for Cryptococcus neoformans and study its functions. Methods Knocking-out and reconstruction the SER5 gene were performed in strain H99, and the function of Ser5 were analyzed by in vitro stress response experiment, melanin induction experiment, capsule induction experiment, urease determination, growth curve determination, and yeast two-hybrid experiment. Results Gene deletion and reconstruction mutants of SER5 were successfully constructed. ser5Δ had no significant effect on the in vitro stress, capsule, urea decomposition and growth of C.neoformans, and it had a significant reduction in the melanin synthesis of C. neoformans.19 genes were screened by yeast two-hybrid experiment that the coding proteins may interact with Ser5. Conclusion Ser5 affected the melanin production of C. neoformans, and it might interact with endoplasmic reticulum protein, protein YOP1, vesicle-associated membrane protein 7, inorganic phosphate transporter pho88, GDP-mannose transporter 1, vesicle transport through interaction with t-SNAREs 1, suggesting that Ser5 might be involved in the synthesis or transport of melanin in C. neoformans.

Key words: Cryptococcus neoformans, serine protease, melanin, function

中图分类号:

R379.5

童双礼, 吴洁, 李莎莎, 桑军军. 新生隐球菌SER5基因的敲除与功能验证研究[J]. 中国真菌学杂志, 2021, 16(5): 335-340.

Tong Shuangli, WU Jie, LI Shasha, SANG Junjun. Deletion and functional studies of SER5 gene in Cryptococcus neoformans[J]. Chinese Journal of Mycology, 2021, 16(5): 335-340.

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