植物中CIPK6、MYB34及MYB51基因的上游開放讀序框(uORF)之探討

Analysis of CIPK6, MYB34 and MYB51 upstream open reading frames in plants

周曉君(Hsiao-Chun Chou)

指導教授 ： 施明哲

共同指導教授 ： 陳荷明(Ho-Ming Chen)

國立臺灣大學/生命科學院/植物科學研究所/碩士(2015年)

https://doi.org/10.6342/NTU.2015.02113

摘要

上游開放讀序框(uORF)為位於5'端非轉譯區中之開放讀序框。過去研究顯示，植物中有些高度保守的上游開放讀序框在高濃度的胺基酸、糖、抗壞血酸或多胺的環境下會導致下游主讀序框 (main ORF)蛋白質轉譯受到抑制。其中有些上游開放讀序框會轉譯出短胜肽鍊並造成核醣體滯留在上游開放讀序框區域，甚至導致此mRNA降解。藉由分析大量RNA降解中間產物，我們發現產生保守胜肽鍊的上游開放讀序框會累積受停滯核糖體保護的RNA降解片段，利用此特徵，我們透過uORF的預測得知在蛋白激酶CIPK6和轉錄因子MYB34及MYB51的5'端非轉譯區具有過去尚未發現之有調控功能的上游開放讀序框。CIPK6參與在植物生長素傳輸和鹽害、乾旱及離層酸反應當中。序列分析顯示CIPK6之上游開放讀序框之胺基酸序列和ATG起始點的周邊序列在不同維管束植物中具高度保守性。且破壞CIPK6 上游開放讀序框會造成下游主讀序框的表現增加。而CIPK6上游開放讀序框對於下游主讀序框的調節，主要藉由影響蛋白轉譯效率，而少部分經由影響RNA量的多寡。MYB34及MYB51與 MYB28、MYB29、MYB122共同參與調控硫代葡萄糖苷生合成途徑。除MYB122上游開放讀序框外，其餘4個MYB皆具有只在十字花科出現，轉譯保守胜肽序列的上游開放讀序框。MYB34及MYB51的上游開放讀序框會抑制下游報導基因活性。在此研究當中，我們證實了由分析RNA降解中間產物找到的上游開放讀序框的調節功能，並闡明一個種系特異性的上游開放讀序框的演化。

關鍵字

阿拉伯芥 ； 十字花科 ； 上游開放讀序框 ； RNA降解體 ； 核糖體滯留 ； CIPK6 ； MYB ； 硫代葡萄糖苷 ； 演化

並列摘要

Highly conserved upstream open reading frame (uORF) in the 5′ untranslated region (UTR) can regulate downstream main ORF translation in response to metabolites. Few uORFs have been shown to stall ribosomes and induce mRNA decay. The genome-wide analysis of RNA degradation intermediates has revealed footprints of stalled ribosomes in conserved peptide uORFs and identified novel regulatory uORFs in genes encoding CBL-interacting protein kinase 6 (CIPK6) and two transcription factors, MYB34 and MYB51. CIPK6 is involved in auxin transport, salt, drought and abscisic acid response. We show that both peptide sequence and ATG context of CIPK6 uORFs are highly conserved in vascular plants and the disruption of CIPK6 uORF enhanced the expression of downstream main ORF. The CIPK6 uORF-mediated regulation is mainly in translation efficiency and lesser in RNA level. MYB34 and MYB51 regulate glucosinolate biosynthesis along with MYB28, MYB29 and MYB122. Except MYB122, they all contain a conserved peptide uORF specific in the Brassicacaeas family. Both MYB51 and MYB34 uORFs repress the downstream reporter activity. Our results validate the regulatory function of uORFs predicted by the analysis of mRNA degradation fragments and unravel the evolution of a lineage-specific uORF.

並列關鍵字

Arabidopsis thaliana ； Brassicacaeas ； PARE ； ribosome stalling ； uORF ； CIPK6 ； MYB ； Glucosinolates (GSL) ； evolution

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