Abstract
Peroxisomes are eukaryotic specific organelles that perform diverse metabolic functions including fatty acid β-oxidation, reactive species metabolism, photorespiration, and responses to stress. However, the potential regulation of these functions by post-translational modifications, including protein phosphorylation, has had limited study. Recently, we identified and catalogued a large number of peroxisomal phosphorylated proteins, implicating the presence of protein kinases in this organelle. Here, we employed available prediction models coupled with sequence conservation analysis to identify 31 protein kinases from the Arabidopsis kinome (all protein kinases) that contain a putative, non-canonical peroxisomal targeting signal type 1 (PTS1). From this, twelve C-terminal domain-PTS1s were demonstrated to be functional in vivo, targeting enhanced yellow fluorescent protein to peroxisomes, increasing the list of presumptive peroxisomal protein kinases to nineteen. Of the twelve protein kinases with functional PTS1s, we obtained full length clones for eight and demonstrated that seven target to peroxisomes in vivo. Screening homozygous mutants of the presumptive nineteen protein kinases revealed one candidate (GPK1) that harbors a sugar-dependence phenotype, suggesting it is involved in regulating peroxisomal fatty acid β-oxidation. These results present new opportunities for investigating the regulation of peroxisome functions.
Highlights
Peroxisomes are eukaryotic specific, single-membraned organelles involved in a host of specialized metabolic events, but are best characterized for their role in fatty acid β-oxidation and photorespiration (Li-Beisson et al, 2013; Kao et al, 2018)
Using the newly developed model “PPero”, we were able to extract seven additional protein kinases that harbor putative peroxisomal targeting signal type 1 (PTS1) (SKD>, 3 with SSL>, SCL>, 2 with SHL>), that were not detected by the positionspecific weight matrices (PWM) or residue interdependence (RI) models (Supplementary Table 1)
We identified four peroxisomal protein phosphatases and predict a higher number of peroxisomal protein kinases based on the genomic ratio of protein phosphatases to kinases (Matre et al, 2009; Kataya et al, 2015b; 2016)
Summary
Peroxisomes are eukaryotic specific, single-membraned organelles involved in a host of specialized metabolic events, but are best characterized for their role in fatty acid β-oxidation and photorespiration (Li-Beisson et al, 2013; Kao et al, 2018). Pre-peroxisomes bud from specific ER domains and become mature peroxisomes through importing nuclear-encoded peroxisomal membrane (PMPs) and matrix proteins. The majority of peroxisomal matrix proteins have PTS1 with a prototype SKL> Bioinformatic prediction tools have been developed primarily for PTS1 identification and have been widely utilized to identify putative peroxisomal candidate proteins (Reumann, 2004; Lingner et al, 2011; Chowdhary et al, 2012; Skoulding et al, 2015; Wang et al, 2017; Reumann and Chowdhary, 2018)
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