蒺藜苜蓿之轉錄因子MtNF-YC6及MtNF-YC11於叢枝菌根菌共生的功能性研究

Functional characterization of Medicago truncatula Nuclear Factor YC6 (MtNF-YC6)and MtNF-YC11 in arbuscular mycorrhizal symbiosis

鄧晨(Chen Deng)

指導教授 ： 楊雯如

共同指導教授 ： 林維怡(Wei-Yi Lin)

國立臺灣大學/生物資源暨農學院/園藝暨景觀學系/碩士(2019年)

https://doi.org/10.6342/NTU201903689

摘要

叢枝菌根菌(Arbuscular mycorrhizal fungi)能促進宿主植物養分的吸收，並提升其對逆境之耐受性，適合作為生物型農藥或肥料來施用。蒺藜苜蓿(Medicago truncatula)的MtNF-YC6和C11為叢枝菌根菌共生保守基因，專一且大量表現於菌根組織。MtNF-YC6和C11屬於核因子Y (Nuclear Factor Y, NF-Y)轉錄因子家族中C亞家族成員，可與A及B亞家族成員組成三聚體以調控目標基因表現。本研究旨在了解這二個轉錄因子於植物與叢枝菌根菌共生關係中所扮演的角色。以酵母菌雙雜合法(yeast two-hybrid)鑑定出與MtNF-YC6和C11產生蛋白質交互作用的MtNF-YA3、A4、A8與MtNF-YB12、B17。以農桿菌滲入法(agroinfiltration) 於邊沁菸草(Nicotiana benthamiana)葉表皮檢測這些成員的次細胞定位，發現MtNF-YB12、B17、C6和C11皆表現在細胞質和細胞核，且共表現MtNF-YB12或B17與MtNF-YC6或C11時表現位置則以細胞核為主。以RNA干擾技術同時降低蒺藜苜蓿根部的MtNF-YC6和C11基因表現量，共得到四株成功降低二個目標基因的轉殖株，但僅有一株根部的叢枝菌根菌拓殖率、叢枝比率及共生標誌基因MtPT4和MtBCP1表現量明顯低於空載體轉殖株。總結本研究結果，目前已鑑定出與MtNF-YC6和C11交互作用之家族成員及其次細胞位置，這些資訊可提供未來探究這些成員在植物與叢枝菌根菌共生關係的線索。

關鍵字

蒺藜苜蓿 ； 叢枝菌根菌共生 ； 核因子Y轉錄因子 ； MtNF-YC6 ； MtNF-YC11

並列摘要

Arbuscular mycorrhizal (AM) fungi which help plants to absorb mineral nutrients from rhizosphere and enhance their tolerance to abiotic and biotic stresses are applied as the biofertilizer for crop production . Medicago truncatula MtNF-YC6 and C11, two highly AM symbiosis-induced genes, are conserved in the genome of AM host plant species. MtNF-YC6 and C11 which belong to C subfamily of Nuclear Factor Y (NF-Y) transcription factor family usually form a trimer with A and B subunits to regulate downstream gene expression. This study aims to investigate the roles of MtNF-YC6 and C11 in AM symbiosis. Through yeast two-hybrid assay, MtNF-YA3, A4, A8, B12 and B17 were identified as potential interacting partners of MtNF-YC6 and C11. The subcellular localization of these proteins were examined in the leaves of Nicotiana benthamiana by using agroinfiltration, and the results suggested that MtNF-YB12, B17, C6 and C11 were localized in the cytosol and the nucleus. Intriguingly, the coexpressing either B12 or B17 with C6 or C11 changed the localization of most proteins to the nucleus. Four MtNF-YC6 and C11 double knockdown Medicago plants were generated by RNA interference techniques. However, only one line showed lower fungal colonization rate, arbuscule formation and the reduction of AM symbiosis marker gene (MtPT4 and MtBCP) expression compared to empty vector controls. In conclusion, identification of the interacting partners of MtNF-YC6 and C11 and their subcellular localization through this study provide clues for studying the roles of MtNF-Y complex in AM symbiosis in the future.

並列關鍵字

Medicago truncatula ； arbuscular mycorrhizal symbiosis ； Nuclear Factor Y ； MtNF-YC6 ； MtNF-YC11

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