Root growth is maintained by the continuous division of cells in the apical meristem. ROOT MERISTEM GROWTH FACTOR 1 (RGF1) is a critical peptide hormone regulating root stem cell niche maintenance. Previous studies discovered that five closely related leucine-rich repeat receptor-like protein kinases (LRR-RLKs), named RGF1 INSENSITIVES (RGIs) or RGF1 RECEPTORS (RGFRs), are able to perceive the RGF1 signal and redundantly control root stem cell niche maintenance. RGF1 regulates root meristem activity mainly via two downstream transcription factors, PLETHORA 1 (PLT1) and PLT2. Regulatory proteins connecting cell surface RGF1-RGI1 and nuclear PLTs, however, were not identified. Here, we report that the mitogen-activated protein (MAP) kinase kinase 4 (MKK4) and MAP kinase 3 (MPK3) were co-immunoprecipitated with RGI1-FLAG after Arabidopsis seedlings were treated with RGF1. Genetic and biochemical assays confirmed that MKK4 and MKK5, and their downstream targets MPK3 and MPK6, are essential RGI-dependent regulators of root meristem development. In addition, we found that the MKK4/MKK5-MPK3/MPK6 module functions downstream of YDA, a MAPKKK. Our results demonstrate that RGF1-RGI1 regulate the expression of PLT1/PLT2 via a YDA-MKK4/MKK5-MPK3/MPK6 signaling cascade.

根部分生组织中细胞的连续分裂维持了根的生长。根瘤生长因子1（RGF1）是调节根干细胞生态位维持的关键肽激素。先前的研究发现，五个紧密相关的富含亮氨酸的重复受体样蛋白激酶（LRR-RLK），分别称为RGF1敏感蛋白（RGI）或RGF1受体（RGFRs），能够感知RGF1信号并冗余控制根干细胞的生态位维持。 。RGF1主要通过两个下游转录因子PLETHORA 1（PLT1）和PLT2调节根分生组织的活性。然而，尚未确定连接细胞表面RGF1-RGI1和核PLT的调控蛋白。在这里，我们报告有丝分裂原激活蛋白（MAP）激酶激酶4（MKK4）和MAP激酶3（MPK3）与RGI1-FLAG共同免疫沉淀后拟南芥幼苗用RGF1处理。遗传和生化分析证实，MKK4和MKK5及其下游靶标MPK3和MPK6是根分生组织发育必不可少的RGI依赖性调节剂。此外，我们发现MKK4 / MKK5-MPK3 / MPK6模块在YDA（MAPKKK）的下游起作用。我们的结果表明RGF1-RGI1调节的表达PLT1 / PLT2经由YDA-MKK4 / MKK5-MPK3 / MPK6信号级联。