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

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.

Cell wall contributes to the stability of plasma membrane nanodomain organization of Arabidopsis thaliana FLOTILLIN2 and HYPERSENSITIVE INDUCED REACTION1 proteins

“…This fast pool of proteins may be in fact the proteins not associated with the PM nanodomains but with the areas of PM where the proteins were not clustered into nanodomains. Therefore we determined coefficient of variation (CV) of fluorescence intensity in linear profile which can be used to assess the fluorescence segregation level (Retzer et al , ), i.e. the degree of aggregation of fluorescence in nanodomains.…”

The PIN-FORMED Auxin Efflux Carriers in Plants

“…These phosphorylation sites are essential for PIN’s polar localization and absent in the same regions of short ones [ 59 ]. In addition to phosphorylation sites, two evolutionarily-conserved cysteine residues (i.e., C39 and C560) on the TD were identified as cis -acting regulators to regulate PIN2 polar localization [ 67 ].…”

Functional analysis of the CDPK-Related Kinase (CRK) family in Arabidopsis thalianal

“…This delayed root geotropic response can be a associated to be a consequence of altered distribution of PIN2 localization in all Atcrk mutants suggesting that auxin transport is impaired to a certain extent in these mutants as was earlier reported in Atcrk5-1 mutant (Rigó et al, 2013). Many auxin transporters are important to maintain the auxin transport in roots, PIN2 auxin efflux transporter is one of the important transporter and plays a role in basipetal auxin transport in Arabidopsis roots (Müller et al, 1998;Retzer et al, 2017).…”