過量表現OsbHLH068水稻轉殖株之根部轉錄體分析

轉錄因子可調控植物的生長、發育與面對逆境的反應。其中basic/helix-loop-helix (bHLH) 轉錄因子為一龐大水稻轉錄因子家族。目前水稻OsbHLHs基因如何參與水稻生長發育及非生物逆境耐受性所知不多。先前本實驗室分析公開微陣列資料庫與本身試驗的微陣列晶片資料結果，篩選出水稻受鹽逆境影響表現的基因 OsbHLH068，並開始對其功能進行了初步探討。研究結果指出於阿拉伯芥過量表現OsbHLH068能促使阿拉伯芥幼苗的根部延長並延遲其開花時間。而過量表現OsbHLH068之水稻則亦會促使水稻幼苗根部的延長，不過其調控的分子機制卻仍不清楚。為瞭解 OsbHLH068 如何影響水稻幼苗根部的生長發育，本試驗藉由RNA定序實驗進行轉錄體分析比較 TNG67 (野生型) 及過量表現水稻轉錄因子 OsbHLH068 轉殖株 (OsbHLH068 OE) 三葉齡幼苗根之基因表現圖譜 (Gene expression profiling) 的變化。結果顯示在兩個過量表現株 OsbHLH068 OE 8 和 OE 12 共同具有467個差異表現基因 (Differentially expressed genes, DEGs)，其中212個為表現量上調基因、255個則是表現量下調基因。經過基因富集分析 (Gene Ontology analysis) 與代謝路徑分析 (Kyoto Encyclopedia of Genes and Genomes pathway analysis)，得知差異表現基因的功能主要參與在逆境刺激反應、脂質的運輸、荷爾蒙訊息傳遞等的反應路徑，其中OsbHLH068可能參與Auxin、Cytokinin、Jasmonate訊息傳遞路徑來影響根長。另外經qPCR試驗證實OsPME1、OsEXPA7受到OsbHLH068正向調控，可能透過促使細胞壁的擴展來影響水稻胚根的延長。綜上所述，OsbHLH068不只參與了水稻非生物逆境耐受性之反應，亦在水稻根系發育上扮演調控角色。

關鍵字

OsbHLH068 ；稉稻；次世代定序；轉錄體表現；代謝路徑分析

並列摘要

Transcription factors regulate plant growth, development and response to various stresses. The basic/helix-loop-helix (bHLH) transcription factor belongs to a large rice transcription factor gene family. However, The functions of OsbHLHs gene involved in rice growth and development are not well studied. From public and laboratory microarray data, we picked up OsbHLH068 which is induced by salt for further study. Functional analysis of OsbHLH068 gene showed that heterologous over-expression OsbHLH068 in Arabidopsis thaliana, results in enhanced root elongation and delayed flowering time. Similar results were also obtained in overexpressed OsbHLH068 transgenic rice which displayed longer root phenotype. However, the regulation of OsbHLH068 in root elongation is not clear. To find out the possible molecular mechanism, we used RNA-sequencing technology and conducted transcriptome analysis of over-expressed transgenic rice root. The results showed that there are 467 differentially expressed genes (DEGs), including 212 up-regulated genes and 255 down-regulated genes. With assistance of Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genome (KEGG), these genes can be categorized into various groups that mainly involved in stress stimulation reaction, lipid transport, and hormone signal transduction. From the results, root growth affected by OsbHLH068 may related to the regulation of Auxin, Cytokinin and Jasmonate signal transduction. Interestingly, RNA-Seq data and qPCR demonstrated that OsPME1 and OsEXPA7 genes were up-regulated in over-expressed OsbHLH068 rice. Taken together, this study showed that OsbHLH068 may not only involved in salinity stress response but also participate in regulating rice root growth.