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標題: | 週期素依賴性蛋白質激酶 Ssn3 調控隱球菌有性生殖 Ssn3, a cyclin‐dependent protein kinase, modulates sexual reproduction in Cryptococcus neoformans |
作者: | Yu-Lin Shi 施佑霖 |
指導教授: | 沈偉強 |
關鍵字: | 隱球菌,Ssn3,Ssn8,雙性有性生殖,單性有性生殖, Cryptococcus neoformans,Ssn3,Ssn8,bisexual mating,unisexual mating, |
出版年 : | 2015 |
學位: | 碩士 |
摘要: | 隱球菌為環境致病性擔子菌,具有兩種交配型 MATa 與 MATα。在氮素源缺乏的情形下,隱球菌可以進行兩種不同有性生殖過程,分別為雙性有性生殖與單性有性生殖。在雙性有性生殖過程中,MATa 與 MATα 菌株會進行細胞融合並產生雙核菌絲,隨後在雙核菌絲的末端發生膨大,接著形成擔子柄與擔孢子,完成有性生殖週期。有趣的是,另外一種類似的有性生殖過程,稱為單性有性生殖,只發生在 MATα 細胞,無需 MATa 細胞的參與;研究顯示,這種獨特的有性生殖方式產生的重組子代,推測與北美近期爆發的隱球菌症有關。隱球菌 Ssn8 蛋白質,為 RNA 聚合酶 II 全酶中的週期素,本實驗室先前藉由逢機遺傳突變篩選得到,並且確認在隱球菌許多的生理過程,包括單性有性生殖,扮演負向調控的角色。為了瞭解單性有性生殖的分子調控網絡,本研究針對另一個會與 Ssn8 蛋白質有交互作用的週期素依賴性蛋白質激酶 Ssn3,進行研究。ssn3 基因突變株,在有性生殖與其他表現型上,與 ssn8 基因突變株相同,其中在單性有性生殖,MATa 與 MATα 的ssn3突變株,皆可觀察到菌絲過度生長的性狀。利用酵母菌雙雜合系統,確認 Ssn3 與 Ssn8 有物理上的交互作用,顯示隱球菌 Ssn8/Ssn3 蛋白質複合體參與單性有性生殖,及其他生理之調控。雖然 ssn3 與 ssn8 突變株有性生殖菌絲生長情形相似,但在雙性有性生殖中,ssn3 突變株產孢時間延遲並且產率下降。利用基因上位性之分析,探討 SSN3 與 SSN8 基因於有性生殖調控路徑中,與 MAT2 及 ZNF2 基因之上下游關係,發現在單性及雙性有性生殖之訊息傳導途徑,SSN3 與 SSN8 基因可能作用於 MAT2 基因下游,ZNF2 基因上游之位置。 Cryptococcus neoformans is an environmental and pathogenic basidiomycete containing two different mating types, MATa and MATα. Under nitrogen starvation conditions, two different sexual reproduction events, bisexual mating and unisexual mating, are observed in C. neoformans. In the processes of bisexual mating, MATa and MATα cells undergo cell fusion to generate dikaryotic filaments. Subsequently, basidia and basidiospores are formed at the tips of dikaryotic filaments to complete the sexual cycle. Interestingly, similar sexual differentiation, so called unisexual mating, can occur only in the MATα cells without MATa cells involved. This unique reproduction event has been implicated to contribute to the generation of recombinant progeny responsible for the recent outbreak of cryptococcosis in North America. C. neoformans Ssn8, a putative cyclin-like component of the RNA polymerase II holoenzyme, was identified in a random mutagenesis screen and shown to play negative roles in diverse physiological processes including unisexual reproduction. To further dissect the molecular networks of unisexual mating in C. neoformans, we here characterized a putative interacting component of Ssn8, the cyclin-dependent protein kinase Ssn3. Mutation of the SSN3 gene reproduced the sexual reproduction and other phenotypes of ssn8 mutants, and hyperfilamentation during unisexual reproduction was observed in the MATα and MATa ssn3 mutant strains. Yeast two-hybrid assay confirmed that Ssn3 physically interacted with Ssn8. These results suggest unisexual reproduction and other physiological processes are regulated by Ssn8/Ssn3 complex in C. neoformans. Although the filamentation phenotypes of ssn3 and ssn8 mutants were similar, delayed and reduced sporulation in bilateral ssn3 mutant cross were observed. To reveal the roles of SSN3 and SSN8 in the signaling pathway of sexual reproduction, we conducted epistasis analysis. The results suggest that SSN3 and SSN8 may function downstream of MAT2 and upstream of ZNF2 during bisexual and unisexual reproduction processes. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/51934 |
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顯示於系所單位: | 植物病理與微生物學系 |
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