Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, Belgium; Center for Plant Systems Biology, VIB, Ghent, Belgium.
Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, Belgium; Center for Plant Systems Biology, VIB, Ghent, Belgium; Department of Biochemistry, Ghent University, Ghent, Belgium; Center for Medical Biotechnology, VIB, Ghent, Belgium.
Mol Cell Proteomics. 2021;20:100040. doi: 10.1074/mcp.RA119.001766. Epub 2021 Jan 7.
The F-box protein MORE AXILLARY GROWTH 2 (MAX2) is a central component in the signaling cascade of strigolactones (SLs) as well as of the smoke-derived karrikins (KARs) and the so far unknown endogenous KAI2 ligand (KL). The two groups of molecules are involved in overlapping and unique developmental processes, and signal-specific outcomes are attributed to perception by the paralogous α/β-hydrolases DWARF14 (D14) for SL and KARRIKIN INSENSITIVE 2/HYPOSENSITIVE TO LIGHT (KAI2/HTL) for KAR/KL. In addition, depending on which receptor is activated, specific members of the SUPPRESSOR OF MAX2 1 (SMAX1)-LIKE (SMXL) family control KAR/KL and SL responses. As proteins that function in the same signal transduction pathway often occur in large protein complexes, we aimed at discovering new players of the MAX2, D14, and KAI2 protein network by tandem affinity purification in Arabidopsis cell cultures. When using MAX2 as a bait, various proteins were copurified, among which were general components of the Skp1-Cullin-F-box complex and members of the CONSTITUTIVE PHOTOMORPHOGENIC 9 signalosome. Here, we report the identification of a novel interactor of MAX2, a type 5 serine/threonine protein phosphatase, designated PHYTOCHROME-ASSOCIATED PROTEIN PHOSPHATASE 5 (PAPP5). Quantitative affinity purification pointed at PAPP5 as being more present in KAI2 rather than in D14 protein complexes. In agreement, mutant analysis suggests that PAPP5 modulates KAR/KL-dependent seed germination under suboptimal conditions and seedling development. In addition, a phosphopeptide enrichment experiment revealed that PAPP5 might dephosphorylate MAX2 in vivo independently of the synthetic SL analog, rac-GR24. Together, by analyzing the protein complexes to which MAX2, D14, and KAI2 belong, we revealed a new MAX2 interactor, PAPP5, that might act through dephosphorylation of MAX2 to control mainly KAR/KL-related phenotypes and, hence, provide another link with the light pathway.
F-box 蛋白 MORE AXILLARY GROWTH 2（MAX2）是独脚金内酯（SLs）以及烟碱衍生的卡瑞琳（KARs）和迄今为止未知的内源性 KAI2 配体（KL）信号级联反应的核心组成部分。这两组分子参与重叠和独特的发育过程，信号特异性结果归因于对 SL 和 KARRIKIN INSENSITIVE 2/HYPOSENSITIVE TO LIGHT（KAI2/HTL）的拟肽α/β-水解酶 DWARF14（D14）的感知。此外，取决于激活的受体，SUPPRESSOR OF MAX2 1（SMAX1）-LIKE（SMXL）家族的特定成员控制 KAR/KL 和 SL 反应。由于在同一信号转导途径中起作用的蛋白质通常存在于大型蛋白质复合物中，我们旨在通过拟南芥细胞培养中的串联亲和纯化来发现 MAX2、D14 和 KAI2 蛋白质网络的新成员。当使用 MAX2 作为诱饵时，共纯化了各种蛋白质，其中包括 Skp1-Cullin-F-box 复合物的一般组成部分和 CONSTITUTIVE PHOTOMORPHOGENIC 9 信号体的成员。在这里，我们报告了 MAX2 的一种新型相互作用蛋白的鉴定，一种 5 型丝氨酸/苏氨酸蛋白磷酸酶，命名为 PHYTOCHROME-ASSOCIATED PROTEIN PHOSPHATASE 5（PAPP5）。定量亲和纯化表明 PAPP5 在 KAI2 中比在 D14 蛋白复合物中更存在。一致地，突变分析表明 PAPP5 可以在非最佳条件下调节 KAR/KL 依赖性种子萌发和幼苗发育。此外，磷酸肽富集实验表明，PAPP5 可能在体内独立于合成的 SL 类似物 rac-GR24 使 MAX2 去磷酸化。总的来说，通过分析 MAX2、D14 和 KAI2 所属的蛋白质复合物，我们揭示了一种新的 MAX2 相互作用蛋白 PAPP5，它可能通过使 MAX2 去磷酸化来主要控制 KAR/KL 相关表型，从而与光途径建立另一个联系。
