拟南芥SRS5基因调控侧根发育分子机制研究

As an essential part of roots system，lateral roots play an important role in plant development and environmental adaptation. However, lateral roots cannot develop infinitely. There are some negative regulators in pericycle to ensure that lateral roots only develop when absolutely required. And auxin can promote lateral root development by removing the negative regulation. However, such negative regulators in a hormone- and signal-independent manner, referred to as ‘intrinsic’ LR regulators remain not being reported. We screened the Arabidopsis mutant rlr1, in which over-expression of SRS5 induced by β-estrogen, resulting in a significant reduction in the number of lateral roots. SRS5 knockout mutant srs5-2 showed more lateral roots than the wild-type, suggesting that SRS5 repressed lateral roots development. We also found that auxin can inhibit the expression of SRS5 in lateral root primordia. And there are some auxin response elements (AuxREs) in the promoter of SRS5 gene, suggesting SRS5 can be directly regulated by IAA/ARF-mediated auxin signaling pathway. Meanwhile, we found that SRS5 directly repressed the expression of LBD16 and LBD29 genes, thereby inhibiting lateral root initiation. On this basis, we plan to study the molecular function of SRS5 in lateral root development. Our study will demonstrate that SRS5, an auxin-inhibited transcription factor, regulates lateral root development via direct repression of LBD genes in Arabidopisis.

侧根是植物根系的重要组成部分，在植物发育和环境适应中起重要作用。然而，侧根并不能无限增殖，中柱鞘细胞中存在负调控因子抑制侧根起始，生长素能解除其抑制作用从而促进侧根发育，目前这类非激素途径成员的侧根发育负调控因子未见报道。我们筛选获得拟南芥突变体rlr1，此突变体中SRS5过量表达，导致侧根数量减少。SRS5功能缺失突变体srs5-2中侧根数量增多，表明SRS5在侧根发育过程中起负调控作用。同时，我们发现生长素抑制SRS5在侧根中的表达。SRS5启动子上具有ARF结合位点，暗示SRS5能够被IAA/ARF介导的生长素信号调控。此外，我们发现SRS5能够调控LBD16/29的表达，抑制侧根起始。我们计划深入分析SRS5调控侧根发育的分子机理，本项目将明确SRS5为侧根起始发育负调控因子，生长素通过其信号途径抑制SRS5基因的表达，解除SRS5对LBD16/29的抑制作用，促进侧根起始发育。

侧根作为植物根系的重要组成部分，在植物发育和环境适应中起重要作用。已有研究显示负责侧根起始的中柱鞘细胞存在负调控因子来抑制侧根起始，而生长素能通过其信号途径解除其抑制作用而促进侧根起始发育，但此类负调控因子未见报道。.本项目通过筛选突变体库，获得拟南芥突变体rlr1。在雌激素诱导下，此突变体中SHI基因家族转录因子SRS5过量表达，导致突变体侧根及侧根原基的数目和密度显著减少。Pro35S:SRS5-GFP株系与rlr1表型一致，而SRS5基因插入失活突变体侧根及侧根原基的数目和密度均较野生型增多，表明转录因子SRS5为侧根发育的负调控因子。利用UAS/GAL4株系，将SRS5基因特异在中柱鞘或早期侧根原基中高量表达，结果显示SRS5既抑制了侧根的起始也抑制了侧根原基发育。LBD16和LBD29是侧根发育中重要的正调控因子。利用LBD16::GUS rlr1和LBD29::GUS rlr1株系，证明SRS5的过表达能显著抑制 LBD16和LBD29的表达。ChIP和EMSA实验证明SRS5能直接结合到LBD16和LBD29启动子上。遗传杂交显示，srs5-2 lbd16， srs5-2 lbd29和srs5-2 lbd16 lbd29分别表现与lbd16，lbd29和lbd16 lbd29一致侧根减少的表型，而35S::LBD16 rlr1和35S::LBD29 rlr1可以恢复rlr1侧根减少的表型，表明SRS5应是通过直接抑制LBD16和LBD29来抑制侧根发育。生长素能解除侧根起始发育负调控因子的抑制作用而促进侧根起始发育。ProSRS5:GUS报告株系显示SRS5基因侧根起始时即有表达，随着侧根原基的发育表达反而减弱。外源IAA处理，SRS5的表达显著降低，而生长素对SRS5基因表达的抑制作用在生长素反应因子ARF7和ARF19的双突变中被解除。ChIP-PCR实验证ARF7和ARF19能与SRS5启动子结合。遗传实验显示生长素能够通过ARF7/19介导的信号途径，抑制SRS5的表达，从而促进侧根的形成。.综上所述，本项目明确证明SRS5为侧根发育的负调控因子。SRS5通过直接结合LBD16和LBD29的启动子降低其表达从而抑制侧根发育。生长素能够通过ARF7和ARF19直接抑制 SRS5的表达，从而解除SRS5的对侧根发育的抑制。