The pseudophosphatase MK-STYX interacts with G3BP and decreases stress granule formation

Abstract

MK-STYX [MAPK (mitogen-activated protein kinase) phospho-serine/threonine/tyrosine-binding protein] is a pseudophosphatase member of the dual-specificity phosphatase subfamily of the PTPs (protein tyrosine phosphatases). MK-STYX is catalytically inactive due to the absence of two amino acids from the signature motif that are essential for phosphatase activity. The nucleophilic cysteine residue and the adjacent histidine residue, which are conserved in all active dual-specificity phosphatases, are replaced by serine and phenylalanine residues respectively in MK-STYX. Mutations to introduce histidine and cysteine residues into the active site of MK-STYX generated an active phosphatase. Using MS, we identified G3BP1 [Ras-GAP (GTPase-activating protein) SH3 (Src homology 3) domain-binding protein-1], a regulator of Ras signalling, as a binding partner of MK-STYX. We observed that G3BP1 bound to native MK-STYX; however, binding to the mutant catalytically active form of MK-STYX was dramatically reduced. G3BP1 is also an RNA-binding protein with endoribonuclease activity that is recruited to ‘stress granules’ after stress stimuli. Stress granules are large subcellular structures that serve as sites of mRNA sorting, in which untranslated mRNAs accumulate. We have shown that expression of MK-STYX inhibited stress granule formation induced either by aresenite or expression of G3BP itself; however, the catalytically active mutant MK-STYX was impaired in its ability to inhibit G3BP-induced stress granule assembly. These results reveal a novel facet of the function of a member of the PTP family, illustrating a role for MK-STYX in regulating the ability of G3BP1 to integrate changes in growth-factor stimulation and environmental stress with the regulation of protein synthesis.

The pseudophosphatase MK‐STYX inhibits stress granule assembly independently of Ser149 phosphorylation of G3BP‐1

“…These data indicate that MK‐STYX inhibits stress granule formation independently of the phosphorylation state of G3BP‐1 at Ser149, and thus suggest that MK‐STYX acts at another point in the signalling pathway. Nevertheless, an interaction between MK‐STYX and G3BP‐1 is supported both by our immunoprecipitation studies , as well as by the observation that the catalytically‐active mutant of MK‐STYX was able to dephosphorylate G3BP‐1 specifically at Ser149 and induce endogenous stress granules. Furthermore, the presence of the active mutant induced the formation of stress granules by the phosphomimetic G3BP‐1 mutant (S149E), as well as larger granules in the presence of S149A‐G3BP.…”

Phylogenetic and genetic linkage between novel atypical dual-specificity phosphatases from non-metazoan organisms

“…The existence of inactive PTPs has been well documented in mammals. These inactive enzymes can interact with signaling proteins and are potential regulators of protein phosphorylation-regulated signaling pathways by heterodimerization with active counterpart PTPs (Wishart and Dixon 1998;Gross et al 2002;Begley and Dixon 2005;Gingras et al 2009;Torii 2009;Hinton et al 2010). In fact, physical interaction among Oca proteins has been detected in high-throughput two hybrid and mass spectrometry approaches (Ito et al 2001;Collins et al 2007).…”

Phosphotyrosine recognition domains: the typical, the atypical and the versatile

“…The arrowhead indicates the catalytic cysteine. Mutations that have experimentally proven to restore catalytic activity are also indicated, for STYX [152], MK-STYX [157] and HD-PTP [159]. EGG4/5, Tensin1 and auxilin are predicted to be devoid of PTP activity owing to the lack of a signature motif residue [121,158].…”