Consensus Substrate Sequence for Protein-tyrosine Phosphatase Receptor Type Z

Abstract

Protein-tyrosine phosphatase receptor type Z (Ptprz) has multiple substrate proteins, including G protein-coupled receptor kinase-interactor 1 (Git1), membrane-associated guanylate kinase, WW and PDZ domain-containing 1 (Magi1), and GTPase-activating protein for Rho GTPase (p190RhoGAP). We have identified a dephosphorylation site at Tyr-1105 of p190RhoGAP; however, the structural determinants employed for substrate recognition of Ptprz have not been fully defined. In the present study, we revealed that Ptprz selectively dephosphorylates Git1 at Tyr-554, and Magi1 at Tyr-373 and Tyr-858 by in vitro and cell-based assays. Of note, the dephosphorylation of the Magi1 Tyr-858 site required PDZ domain-mediated interaction between Magi1 and Ptprz in the cellular context. Alignment of the primary sequences surrounding the target phosphotyrosine residue in these three substrates showed considerable similarity, suggesting a consensus motif for recognition by Ptprz. We then estimated the contribution of surrounding individual amino acid side chains to the catalytic efficiency by using fluorescent peptides based on the Git1 Tyr-554 sequence in vitro. The typical substrate motif for the catalytic domain of Ptprz was deduced to be Glu/Asp-Glu/Asp-Glu/Asp-Xaa-Ile/ Val-Tyr(P)-Xaa (Xaa is not an acidic residue). Intriguingly, a G854D substitution of the Magi1 Tyr-858 site matching better to the motif sequence turned this site to be susceptible to dephosphorylation by Ptprz independent of the PDZ domainmediated interaction in cells. Furthermore, we found by database screening that the substrate motif is present in several proteins, including paxillin at Tyr-118, its major phosphorylation site. Expectedly, we verified that Ptprz efficiently dephosphorylates paxillin at this site in cells. Our study thus provides key insights into the molecular basis for the substrate recognition of Ptprz.Protein-tyrosine phosphorylation is a dynamic process governed by the balanced actions of protein-tyrosine kinases (PTKs), 2 and protein-tyrosine phosphatases (PTPs), and critical to the regulation of numerous physiological processes (for review, see Ref. 1). The specificity of the signal transduction depends on the ability of each PTK or PTP to phosphorylate or dephosphorylate precisely particular sites on specific substrates, respectively. Elucidation of the specificity for individual PTPs has been an important subject of investigation; however, even the identification of PTP substrates is still methodologically difficult. To our knowledge, substrate specificity of PTPs has been characterized only for PTP1B (2).Receptor-like PTPs (RPTPs) are a structurally and functionally diverse family of enzymes comprised of eight subfamilies. PTP receptor type Z (Ptprz, also called PTP or RPTP␤) is a RPTP classified in the R5 subfamily together with Ptprg (PTP␥). Three isoforms of Ptprz are generated by alternative splicing from a single Ptprz gene: the two transmembrane isoforms Ptprz-A and Ptprz-B and the secretory isoform Ptprz-S (also known as phosphacan...

Role of Chondroitin Sulfate (CS) Modification in the Regulation of Protein-tyrosine Phosphatase Receptor Type Z (PTPRZ) Activity

“…The extracellular regions of PTPRZ-A/B are highly modified with CS chains in brain tissues and OPCs. In contrast, a lower CS modification was observed when PTPRZ-B was exogenously expressed in BHK-21 (the present study) and HEK293T cells (54) and in endogenous PTPRZ-B expressed in stomach tissue (55) and C6 glioblastoma cells (56). PTPRZ-A and PTPRZ-S contain a long serine, glycine-rich domain extracellularly that is thought to be the CS attachment region.…”

Intracellular and extracellular domains of protein tyrosine phosphatase PTPRZ-B differentially regulate glioma cell growth and motility

“…Here, migration impairment in E98-shPTPRZ1 cells co-incided with reduced MET activity. For a candidate substrate, rather increased phosphorylation levels are expected upon knock-down of PTPRZ, and thus far MET does not meet PTPRZ substrate criteria [44]. Together with ErbB and PDGFR family members, MET represents one of the oncogenic drivers in glioma tumor biology [7].…”

Protein tyrosine phosphatase receptor-like genes are frequently hypermethylated in sporadic colorectal cancer

“…PTPRZ1 has also been reported to play an important role in the functioning of neurons (15). There are no reports on its role as a tumor suppressor or its significance for cell homeostasis.…”

“…inducing gastric ulcers (15). PTPRZ1 has also been reported to play an important role in the functioning of neurons (15).…”