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標題: 以elf18、flg22及VirB2胜肽探討阿拉伯芥RAB8B及8D蛋白質於農桿菌感染過程及植物防禦機制之功能分析
Functional characterization of Arabidopsis thaliana RAB8B and 8D proteins in Agrobacterium tumefaciens infection and plant defense response via elf18, flg22, and VirB2 peptide treatments
作者: 簡珮如
Pei-Ru Chien
關鍵字: 阿拉伯芥;農桿菌;病原菌胜肽;Arabidopsis;Agrobacterium;RAB;elf18;flg22;VirB2
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Bacterial disease resistance in Arabidopsis through flagellin perception. Nature 428, 764-767.
植物的細胞膜表面會有PRR (pattern recognition receptors)受體,能辨認病原菌相關分子圖譜(pathogen/microbe-associated molecular pattern、PAMP/MAMP),例如:植物EFR (EF-Tu Receptor)可識別農桿菌之延伸因子(elongation factor Tu、EF-Tu)的elf18胜肽,或是FLS2 (FLAGELLIN SENSITIVE 2)可以辨認番茄細菌性斑點病菌(Pseudomonas syringae pv. tomato DC3000)之鞭毛蛋白質的flg22胜肽等,會促使植物引發免疫反應,以抵抗病原菌入侵。另已知農桿菌中組成T線毛之主要致病蛋白質VirB2會與AtRAB8B與AtRAB8D結合,且AtRAB8B與AtRAB8D也會與番茄細菌性斑點病菌之致病蛋白質AvrPto結合。其中RAB8B、8D為植物small GTPase RAB8/RABE家族成員,主要位於高基氏體且協助囊泡的形成與運輸。因此為了瞭解AtRAB8B與AtRAB8D在植物防禦反應中的功能,本研究分離獲得rab8b與rab8d突變株,此二類突變株皆不易受農桿菌感染。另外以elf18、flg22及VirB2胜肽預先處理野生株6小時後,再施以農桿菌感染,由實驗結果得知與未處理組相比,野生株感染率皆下降,可能因這三類胜肽可誘發植物的防禦反應所致。而於相同實驗處理下,rab8b與rab8d突變株的感染率則會高於野生株,可能因突變株受e1f18和flg22誘發植株的防禦能力較野生株差所致。本研究再以Real-time PCR方式檢測e1f18、flg22及VirB2胜肽誘導植物後,植株中防禦相關基因的表現量,結果顯示突變株中FRK1、WRKY22、WRKY29、MPK6基因表現量皆較野生株低,顯示當RAB8B與RAB8D基因突變時,可能會使植株的防禦反應異常,導致突變株對病原菌感染的效率與野生株不同。綜合以上結果得知AtRAB8B與AtRAB8D可能透過植物內膜系統的運輸,協助調控防禦反應訊息傳遞的過程,進而影響植物防禦相關基因的表現。

Plants detect presences of various pathogens through recognizing pathogen-associated molecular pattern (PAMPs) or microbe-associated molecular patterns (MAMPs) by pattern recognition receptors (PRRs). The EF-Tu receptor (EFR), one of the PRRs, can recognize the elf18 peptide of the elongation factor Tu from Agrobacterium tumefaciens. Another PRR, the FLAGELLIN SENSITIVE 2 (FLS2), recognizes the flg22 peptide of the flagellin protein from Pseudomonas syringae pv. tomato DC3000. These PAMPs/MAMPs can trigger plant defense response to counteract pathogen infections. The Arabidopsis thaliana RAB8B and RAB8D proteins can interact with the VirB2, the major component of the A. tumefaciens T-pilus. Additionally, the AtRAB8B and 8D proteins interact with a Pst DC3000 effector, the AvrPto protein. The RAB8B and 8D proteins, members of the small GTPase RAB8/RABE family, are localized in the Golgi apparatus and regulate vacuolar formations and intracellular vesicle transports. To understand functions of the RAB8B and 8D during plant defense response, we have identified Arabidopsis rab8b and rab8d T-DNA insertion mutants. The rab8b and rab8d mutants were resistant to root-based A. tumefaciens infections. The wild-type, rab8b and rab8d mutants were then pretreated with the elf18, flg22, or VirB2 peptide for 6 hours before seedling-based A. tumefaciens infections, the infection rates of these plants were decreased in comparison to untreated plants. These data suggest that pretreatments of the elf18, flg22, or VirB2 peptide may induce plant defense responses and help plants to hinder bacteria infections. The rab8b and rab8d mutants became more susceptible to A. tumefaciens infections compared to wild-type plants after pretreatments of the elf18 or flg22, suggesting that induced plant defense responses might be compromised in these mutants. The defense-related gene expressions, including FRK1, WRKY22, WRKY29, and MPK6 genes, were less induced in rab8b and rab8d mutants after elf18, flg22, or VirB2 peptide treatments in comparison to wild-type plants. These data support that lower expressions of the RAB8B or RAB8D genes in mutants might affect plant defense response and therefore influence bacteria infection rates. In conclusion, the AtRAB8B and AtRAB8D may affect plant defense response gene expressions via participating endomembrane trafficking and influencing signal transduction pathways of plant defense responses in plant cells.
Rights: 同意授權瀏覽/列印電子全文服務,2022-02-11起公開。
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