Evolutionary Conserved Cysteines Function as cis-Acting Regulators of Arabidopsis PIN-FORMED 2 Distribution.

Katarzyna Retzer,Nataliia Konstantinova,Roman Skokan,Martina Laňková,Stanislav Vosolsobě,Christian Luschnig,Jozef Lacek,Kateřina Malínská,Richard Napier,Charo Del Genio,Eva Zažímalová,Jan Petrášek

doi:10.3390/ijms18112274

Abstract

Coordination of plant development requires modulation of growth responses that are under control of the phytohormone auxin. PIN-FORMED plasma membrane proteins, involved in intercellular transport of the growth regulator, are key to the transmission of such auxin signals and subject to multilevel surveillance mechanisms, including reversible post-translational modifications. Apart from well-studied PIN protein modifications, namely phosphorylation and ubiquitylation, no further post-translational modifications have been described so far. Here, we focused on root-specific Arabidopsis PIN2 and explored functional implications of two evolutionary conserved cysteines, by a combination of in silico and molecular approaches. PIN2 sequence alignments and modeling predictions indicated that both cysteines are facing the cytoplasm and therefore would be accessible to redox status-controlled modifications. Notably, mutant pin2C−A alleles retained functionality, demonstrated by their ability to almost completely rescue defects of a pin2 null allele, whereas high resolution analysis of pin2C−A localization revealed increased intracellular accumulation, and altered protein distribution within plasma membrane micro-domains. The observed effects of cysteine replacements on root growth and PIN2 localization are consistent with a model in which redox status-dependent cysteine modifications participate in the regulation of PIN2 mobility, thereby fine-tuning polar auxin transport.

Highlights

Auxin, a versatile plant growth regulator, is involved in a multitude of developmental processes [1,2,3]
To address the hypothesis that cysteines do function as cis-acting regulators of PIN protein function, we reasoned that such residues should exhibit a high degree of conservation within the PIN family
This variability ranged from eight residues found in the PIN5 ORF (Open Reading Frame) to only two cysteines found in PIN2 (Figure 1A)

Summary

Introduction

A versatile plant growth regulator, is involved in a multitude of developmental processes [1,2,3]. This versatility is largely dependent on a very flexible molecular machinery, mediating directional transport of the phytohormone throughout the entire organism [4,5,6]. Plasma membrane localized PIN-FORMED (PIN) proteins, in particular, have been connected to the cellular efflux of the growth regulator, a critical step that defines directionality and rates of polar auxin transport and requires tight regulation [4,7]. Plasma membrane-resident PIN proteins are subject to lateral diffusion processes, which is eventually followed by clathrin-dependent endocytic sorting to the TGN. PINs appear to be either rerouted to the plasma membrane, or sorted towards late endosomes en route to the lytic vacuole [4,7]

Methods

Results

Conclusion