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標題: | 阿拉伯芥SAURs的突變降低花粉管生長所必需
之有效轉錄產物的轉譯 Mutation of Arabidopsis SAURs impairs the efficient translation of transcripts essential for pollen tube growth |
作者: | Siou-Luan He 何秀鑾 |
指導教授: | 趙光裕 |
關鍵字: | SAUR,核醣體組裝,肌動蛋白細胞骨架,果膠壁, SAUR,ribosome assembly,actin cytoskeleton,pectin wall, |
出版年 : | 2018 |
學位: | 博士 |
摘要: | 花粉管順利生長對於雙重受精是重要的。了解花粉管基因的生物功能將為這一關鍵過程中的調控機制提供新的見解。根據先前轉譯體學的研究,獲得經授粉作用誘導大量表現的基因SAUR62和SAUR75。SAUR62和SAUR75蛋白質位於花粉管內的細胞核。同質結合突變株saur62/-、saur75/-和SAUR62/75 RNAi轉殖株的果莢內有許多不孕的種子。saur62、saur75、RNAi的花粉活力是正常的,但是其體外、體內花粉管生長是有缺陷並且花粉管呈現分支的型態。轉植株SAUR62/75-GFP花其免疫沉澱實驗顯示核醣體蛋白質RPL12家族可能是SAUR62和SAUR75交互作用的蛋白。藉由酵母菌雙雜合系統和雙分子螢光互補分析實驗證明SAUR62和SAUR75可以和核糖體蛋白質RPL12交互作用。多聚核醣體圖譜實驗顯示saur62、saur75、rpl12c和RNAi的花中80S核醣體減少,推測SAUR62和SAUR75在核醣體組裝的角色。為了闡明SAUR62/75在轉譯作用中的角色,通過iTRAQ分析WT和RNAi花的總蛋白,顯示參與花粉管壁生成和肌動蛋白骨架的蛋白質表現量顯著降低,這對於花粉管延長其管壁的彈性是關鍵的。 實際上,RNAi花粉管顯示異常脫酯化和酯化的果膠和F-肌動蛋白質的分布。 我們的研究證明SAUR62 /75及其交互作用蛋白質RPL12家族成員在核醣體組裝中的角色,對於有效的花粉管伸長和隨後的受精作用是至關重要。 Successful pollen tube elongation is critical for double fertilization. Understanding the biological functions of pollen tube genes will provide new insights into the regulatory machinery underlying this crucial process. Previous translatomic study revealed two Arabidopsis Small Auxin Up RNA genes, SAUR62 and SAUR75, with expression upregulated by pollination. Both SAUR62 and SAUR75 mainly localized in pollen tube nuclei, and siliques of homozygous saur62 (saur62/-), saur75 (saur75/-), and the SAUR62/75 RNAi knockdown line had many aborted seeds. Pollen viability of these mutants and RNAi lines was normal, but in vitro and in vivo pollen tube growth was defective, with branching phenotypes. Immunoprecipitation with transgenic SAUR62/75-GFP flowers revealed ribosomal protein RPL12 family members as their potential partners, and their individual interactions were further confirmed by yeast two-hybrid (Y2H) and bimolecular fluorescence complementation assay (BiFC). Polysome profiling showed reduced 80S ribosome abundance in homozygous saur62, saur75, rpl12c and SAUR62/75 RNAi flowers, which suggests roles for SAUR62/75 in ribosome assembly. To clarify the roles in translation, total proteins of wild-type (WT) and RNAi flowers were analyzed by iTRAQ, which revealed significantly reduced expression of factors participating in pollen tube wall biogenesis and F-actin dynamics, critical for the elastic properties of tube elongation. Indeed, RNAi pollen tubes showed dislocalization of de-esterified and esterified pectins and F-actin organization. Here, we demonstrate the biological roles of SAUR62/75 and their partners, RPL12 family members, as critical in ribosome assembly for efficient pollen tube elongation and the following fertilization. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/22127 |
DOI: | 10.6342/NTU201802103 |
全文授權: | 未授權 |
顯示於系所單位: | 植物科學研究所 |
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