利用染色質免疫沉澱法找尋水稻轉錄因子ERF106MZ下游調控基因

Abstract

APETALA2/ethylene-responsive factors轉錄因子 (AP2/ERFs) 除了參與調控植物生長發育與逆境耐受性外，也與多種植物荷爾蒙作用有關。本實驗室過去的研究發現OsERF106MZ受鹽分誘導，且過量表達OsERF106MZ (OsERF106MZ-OE) 會導致水稻幼苗地上部生長遲緩、降低鹽逆境耐受性。透過DNA微陣列分析比較OsERF106MZ-OE和WT的轉錄體，我們鑑定出多個與逆境反應有關的差異表現基因。進一步進行染色質免疫沉澱定序 (Chromatin immunoprecipitation sequencing, ChIP-seq)，並與DNA微陣列數據結合後，最終我們從50個潛在的OsERF106MZ下游調控基因中選取OsARF11 (Auxin response factor 11) 及 OsPOX22.3 (Peroxidase 22.3) 進行後續分析。結果顯示在OsERF106MZ-OE背景下，不論正常環境或鹽逆境，兩基因的表現量均受到抑制。此外，生長分析結果則顯示OsERF106MZ-OE降低了水稻幼苗對生長素的敏感性。ChIP-qPCR結果顯示，OsERF106MZ可分別與OsARF11之5’ UTR以及OsPOX22.3啟動子區域的GCC box進行結合。綜合上述，這些結果說明OsERF106MZ調控的下游基因中包含了OsARF11與OsPOX22.3，並且為OsERF106MZ調控水稻株高與鹽逆境耐受性之機制提供新的線索。

The APETALA2/ethylene response factor (AP2/ERF) family transcription factors are involved in the regulation of plant growth, stress tolerance, and signaling of phytohormones. Our previous study found that overexpression of the salt inducible OsERF106MZ (OsERF106MZ-OE) can cause growth retardation in the shoots of rice seedlings and reduce tolerance to salt stress. To further dissect the molecular mechanism, comparative analysis of DNA microarray and Chromatin immunoprecipitation sequencing (ChIP-seq) were conducted between OsERF106MZ-OE and WT. By combining differentially expressed genes with ChIP‐seq data, 50 downstream target genes of OsERF106MZ were identified. Two candidate genes, OsARF11 (Auxin response factor 11), and OsPOX22.3 (Peroxidase 22.3), were selected for follow-up analysis. The results showed that under the OsERF106MZ-OE background, the expression levels of both genes were reduced regardless of the normal environment and salt stress. Furthermore, growth analysis results showed that OsERF106MZ-OE reduced the sensitivity of rice to auxin. The ChIP-qPCR analysis showed that OsERF106MZ can bind to the GCC box in the 5’ UTR of OsARF11 and the promoter region of OsPOX22.3, respectively. The results indicate that OsARF11 and OsPOX22.3 are the downstream regulatory genes of OsERF106MZ. These findings offer new clues for studying the molecular mechanism of OsERF106MZ-regulated plant height and salt stress tolerance in rice.