Choi Hee-Seung, Seo Minji, Cho Hyung-Taeg
Department of Biological Sciences, Seoul National University, Seoul, South Korea.
Front Plant Sci. 2018 Mar 21;9:372. doi: 10.3389/fpls.2018.00372. eCollection 2018.
Auxin signaling is finalized by activator auxin response factors (aARFs) that are released from Auxin/Indole-3-Acetic Acid (Aux/IAA) repressors and directly activate auxin-responsive genes. However, it remains to be answered how repressor ARFs (rARFs) exert their repression function. In this study, we assessed the molecular and biological functions of two putative co-repressor-binding motifs (EAR and RLFGI) of ARF2 (a rARF) in . In the yeast two-hybrid assay, the EAR mutation moderately and the RLFGI mutation, or both motifs, almost completely disrupted the interaction between the co-repressor TOPLESS (TPL) and the repressive motifs-containing middle domain (MD) of ARF2. The ARF2-MD interacted not only with TPL but also with TPL homologs (TPRs). Root hair-specific overexpression of rARFs (ARF1-4, 9-11, and 16) considerably inhibited root hair growth, suggesting that rARFs generally function as repressors in the auxin-responsive root hair single cell. Individual mutation of the ARF2 EAR or RLFGI motif slightly and both mutations greatly compromised ARF2-mediated inhibition of root hair growth and auxin-responsive gene expression. In addition, flowering time and seed size, two representative mutant phenotypes, were examined to assess the function of the repressive motifs in mutant-complementation experiments. ARF2-mediated inhibition of flowering and seed growth was suppressed considerably by the individual mutation of EAR or RLFGI and almost completely by both mutations. These results suggest that EAR and RLFGI work together as major repressive motifs for ARF2 to recruit TPL/TPR co-repressors and to exhibit its repressive biological functions.
生长素信号传导由激活型生长素响应因子（aARFs）完成，这些因子从生长素/吲哚-3-乙酸（Aux/IAA）阻遏物中释放出来，并直接激活生长素响应基因。然而，阻遏型ARFs（rARFs）如何发挥其抑制功能仍有待解答。在本研究中，我们评估了ARF2（一种rARF）的两个假定的共阻遏物结合基序（EAR和RLFGI）在[具体内容缺失]中的分子和生物学功能。在酵母双杂交试验中，EAR突变适度破坏，而RLFGI突变或两个基序同时突变几乎完全破坏了共阻遏物TOPLESS（TPL）与ARF2含抑制基序的中间结构域（MD）之间的相互作用。ARF2-MD不仅与TPL相互作用，还与TPL同源物（TPRs）相互作用。根毛特异性过表达rARFs（ARF1-4、9-11和16）显著抑制根毛生长，表明rARFs在生长素响应的根毛单细胞中通常起阻遏作用。ARF2的EAR或RLFGI基序单独突变对根毛生长和生长素响应基因表达的抑制作用略有影响，而两个基序同时突变则大大削弱了这种抑制作用。此外，在突变体互补实验中，检测了开花时间和种子大小这两个代表性的突变体表型，以评估抑制基序的功能。EAR或RLFGI单独突变可显著抑制ARF2介导的开花和种子生长抑制，而两个基序同时突变几乎完全抑制。这些结果表明，EAR和RLFGI共同作为ARF2的主要抑制基序，招募TPL/TPR共阻遏物并发挥其抑制生物学功能。
