Unraveling the MAX2 Protein Network in Arabidopsis thaliana: Identification of the Protein Phosphatase PAPP5 as a Novel MAX2 Interactor

Anse Jacobs,Ive De Smet,Annick De Keyser,Stephen Depuydt,François-Didier Boyer,Kris Gevaert,Geert Persiau,Alan Walton,Carolien De Cuyper,Lukas Braem,Sylwia Struk,Sofie Goormachtig,Geert De Jaeger,Lam Dai Vu,Dominique Eeckhout

doi:10.1074/mcp.ra119.001766

Abstract

AbstractThe 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.

Highlights

IntroductionF-box proteins represent one of the largest and most heterogeneous superfamilies in plants that modulate crucial processes from embryogenesis to seedling development, hormone signaling, and defense pathways [1]
tandem affinity purification (TAP) revealed PHYTOCHROME-ASSOCIATED PROTEIN PHOSPHATASE 5 (PAPP5) as a novel interactor of MORE AXILLARY GROWTH 2 (MAX2). PAPP5 might act through dephosphorylation of MAX2. PAPP5 modulates KARRIKIN INSENSITIVE 2 (KAI2)-controlled responses in seeds and young seedlings. PAPP5 might be another link between KAR/KAI2 ligand (KL) signaling and light
Arabidopsis cell suspension cultures were chosen for TAP experiments owing to the high protein yield and the possibility to carry out trigger-based studies [42]

Summary

Introduction

F-box proteins represent one of the largest and most heterogeneous superfamilies in plants that modulate crucial processes from embryogenesis to seedling development, hormone signaling, and defense pathways [1] They confer substrate specificity of the S-phase kinase-associated protein 1 (Skp1), Cullin-1/CDC53, and F-box protein (SCF) E3 ubiquitin ligase complex that selects proteins for ubiquitination to mark them for proteolysis by the 26S proteasome [2, 3]. One of the Arabidopsis thaliana F-box proteins, MORE AXILLARY GROWTH 2 (MAX2) is a central component of two distinct signaling pathways, one that perceives the plant hormones strigolactones (SLs) and one that senses the smoke-derived karrikins (KARs) [4, 5].

Objectives

Results

Conclusion