Function Of Tyrosine Phosphatases Research Articles

Pharmacological Targeting of the RAGE-NFκB Signalling Axis Impedes Monocyte Activation under Diabetic Conditions through the Repression of SHP-2 Tyrosine Phosphatase Function.

Monocytes play a vital role in the development of cardiovascular diseases. Type 2 diabetes mellitus (T2DM) is a major CVD risk factor, and T2DM-induced aberrant activation and enhanced migration of monocytes is a vital pathomechanism that leads to atherogenesis. We recently reported the upregulation of SHP-2 phosphatase expression in mediating the VEGF resistance of T2DM patient-derived monocytes or methylglyoxal- (MG, a glucose metabolite and advanced glycation end product (AGE) precursor) treated monocytes. However, the exact mechanisms leading to SHP-2 upregulation in hyperglycemic monocytes are unknown. Since inflammation and accumulation of AGEs is a hallmark of T2DM, we hypothesise that inflammation and AGE-RAGE (Receptor-for-AGEs) signalling drive SHP-2 expression in monocytes and blockade of these pathways will repress SHP-2 function. Indeed, monocytes from T2DM patients revealed an elevated SHP-2 expression. Under normoglycemic conditions, the serum from T2DM patients strongly induced SHP-2 expression, indicating that the T2DM serum contains critical factors that directly regulate SHP-2 expression. Activation of pro-inflammatory TNFα signalling cascade drove SHP-2 expression in monocytes. In line with this, linear regression analysis revealed a significant positive correlation between TNFα expression and SHP-2 transcript levels in T2DM monocytes. Monocytes exposed to MG or AGE mimetic AGE-BSA, revealed an elevated SHP-2 expression and co-treatment with an NFκB inhibitor or genetic inhibition of p65 reversed it. The pharmacological inhibition of RAGE was sufficient to block MG- or AGE-BSA-induced SHP-2 expression and activity. Confirming the importance of RAGE-NFκB signalling in regulating SHP-2 expression, the elevated binding of NFκB to the SHP-2 promoter-induced by MG or AGE-BSA-was reversed by RAGE and NFκB inhibition. Besides, we detected elevated RAGE levels in human and murine T2DM monocytes and monocytes exposed to MG or AGE-BSA. Importantly, MG and AGE-BSA treatment of non-T2DM monocytes phenocopied the aberrant pro-migratory phenotype of T2DM monocytes, which was reversed entirely by either SHP-2- or RAGE inhibition. In conclusion, these findings suggest a new therapeutic approach to prevent accelerated atherosclerosis in T2DM patients since inhibiting the RAGE-NFκB-SHP-2 axis impeded the T2DM-driven, SHP-2-dependent monocyte activation.

Mutations that impair Eyes absent tyrosine phosphatase activity in vitro reduce robustness of retinal determination gene network output in Drosophila.

A limited collection of signaling networks and transcriptional effectors directs the full spectrum of cellular behaviors that comprise development. One mechanism to diversify regulatory potential is to combine multiple biochemical activities into the same protein. Exemplifying this principle of modularity, Eyes absent (Eya), originally identified as a transcriptional co-activator within the retinal determination gene network (RDGN), also harbors tyrosine and threonine phosphatase activities. Although mounting evidence argues for the importance of Eya’s phosphatase activities to mammalian biology, genetic rescue experiments in Drosophila have shown that the tyrosine phosphatase function is dispensable for normal development. In this study, we repeated these rescue experiments in genetically sensitized backgrounds in which the dose of one or more RDGN factor was reduced. Heterozygosity for sine oculis or dachshund, both core RDGN members, compromised the ability of phosphatase-dead eya, but not of the control wild type eya transgene, to rescue the retinal defects and reduced viability associated with eya loss. We speculate that Eya’s tyrosine phosphatase activity, although non-essential, confers robustness to RDGN output.

Protein Tyrosine Phosphatases in Systemic Sclerosis: Potential Pathogenic Players and Therapeutic Targets.

The pathogenesis of systemic sclerosis depends on a complex interplay between autoimmunity, vasculopathy, and fibrosis. Reversible phosphorylation on tyrosine residues, in response to growth factors and other stimuli, critically regulates each one of these three key pathogenic processes. Protein tyrosine kinases, the enzymes that catalyze addition of phosphate to tyrosine residues, are known players in systemic sclerosis, and tyrosine kinase inhibitors are undergoing clinical trials for treatment of this disease. Until recently, the role of tyrosine phosphatases-the enzymes that counteract the action of tyrosine kinases by removing phosphate from tyrosine residues-in systemic sclerosis has remained largely unknown. Here, we review the function of tyrosine phosphatases in pathways relevant to the pathogenesis of systemic sclerosis and their potential promise as therapeutic targets to halt progression of this debilitating rheumatic disease. Protein tyrosine phosphatases are emerging as important regulators of a multitude of signaling pathways and undergoing validation as molecular targets for cancer and other common diseases. Recent advances in drug discovery are paving the ways to develop new classes of tyrosine phosphatase modulators to treat human diseases. Although so far only few reports have focused on tyrosine phosphatases in systemic sclerosis, these enzymes play a role in multiple pathways relevant to disease pathogenesis. Further studies in this field are warranted to explore the potential of tyrosine phosphatases as drug targets for systemic sclerosis.

Multiple Functions of the Eya Phosphotyrosine Phosphatase.

Eyes absent (Eya), a protein conserved from plants to humans and best characterized as a transcriptional coactivator, is also the prototype for a novel class of eukaryotic aspartyl protein tyrosine phosphatases. This minireview discusses recent breakthroughs in elucidating the substrates and cellular events regulated by Eya's tyrosine phosphatase function and highlights some of the complexities, new questions, and surprises that have emerged from efforts to understand how Eya's unusual multifunctionality influences developmental regulation and signaling.

New and Unexpected Biological Functions for the Src-Homology 2 Domain-Containing Phosphatase SHP-2 in the Gastrointestinal Tract.

SHP-2 is a tyrosine phosphatase expressed in most embryonic and adult tissues. SHP-2 regulates many cellular functions including growth, differentiation, migration, and survival. Genetic and biochemical evidence show that SHP-2 is required for rat sarcoma viral oncogene/extracellular signal-regulated kinases mitogen-activated protein kinase pathway activation by most tyrosine kinase receptors, as well as by G-protein–coupled and cytokine receptors. In addition, SHP-2 can regulate the Janus kinase/signal transducers and activators of transcription, nuclear factor-κB, phosphatidyl-inositol 3-kinase/Akt, RhoA, Hippo, and Wnt/β-catenin signaling pathways. Emerging evidence has shown that SHP-2 dysfunction represents a key factor in the pathogenesis of gastrointestinal diseases, in particular in chronic inflammation and cancer. Variations within the gene locus encoding SHP-2 have been associated with increased susceptibility to develop ulcerative colitis and gastric atrophy. Furthermore, mice with conditional deletion of SHP-2 in intestinal epithelial cells rapidly develop severe colitis. Similarly, hepatocyte-specific deletion of SHP-2 induces hepatic inflammation, resulting in regenerative hyperplasia and development of tumors in aged mice. However, the SHP-2 gene initially was suggested to be a proto-oncogene because activating mutations of this gene were found in pediatric leukemias and certain forms of liver and colon cancers. Moreover, SHP-2 expression is up-regulated in gastric and hepatocellular cancers. Notably, SHP-2 functions downstream of cytotoxin-associated antigen A (CagA), the major virulence factor of Helicobacter pylori, and is associated with increased risks of gastric cancer. Further compounding this complexity, most recent findings suggest that SHP-2 also coordinates carbohydrate, lipid, and bile acid synthesis in the liver and pancreas. This review aims to summarize current knowledge and recent data regarding the biological functions of SHP-2 in the gastrointestinal tract.

PIK3CA and PTEN mutations in adenoid cystic carcinoma of the breast metastatic to kidney

Adenoid cystic carcinoma (ACC) of the breast rarely metastasizes and has been associated with excellent prognosis. We describe a patient with renal metastasis of primary breast ACC 5 years after the mastectomy. A detailed molecular genetic analysis of the primary and metastatic tumors demonstrated somatic mutations in 2 well-known cancer genes associated with regulation of PI3K/AKT signaling pathway: (1) PIK3CA, which encodes the catalytic alpha subunit of the phosphoinositide-3-kinase, and (2) PTEN, which encodes phosphatase and tensin homolog. The mutation identified in PIK3CA (Ex1+169 A>C) predicts an amino acid change from isoleucine to methionine at codon 31 (I31M) and resides in the p85-binding domain of exon 1. The mutation identified in PTEN (IVS4-3 C>T) resides in intron 4 near the splice acceptor site of exon 5 and was associated with an aberrant PTEN transcript lacking exon 5, which is necessary for protein tyrosine phosphatase function and tumor suppressor properties of PTEN. Increased promoter methylation of PTEN was present in renal metastasis, coinciding with the decrease in the level of normal PTEN transcript. These coexistent mutations/epigenetic inactivations in PI3K/AKT pathway may be responsible for the unusually aggressive course of ACC.

Protein tyrosine phosphatase function: the substrate perspective

It is now well established that the members of the PTP (protein tyrosine phosphatase) superfamily play critical roles in fundamental biological processes. Although there has been much progress in defining the function of PTPs, the task of identifying substrates for these enzymes still presents a challenge. Many PTPs have yet to have their physiological substrates identified. The focus of this review will be on the current state of knowledge of PTP substrates and the approaches used to identify them. We propose experimental criteria that should be satisfied in order to rigorously assign PTP substrates as bona fide. Finally, the progress that has been made in defining the biological roles of PTPs through the identification of their substrates will be discussed.

Radiotherapy-induced signal transduction

Exposure of tumor cells to ionizing radiation causes compensatory activation of multiple intracellular survival signaling pathways to maintain viability. In human carcinoma cells, radiation exposure caused an initial rapid inhibition of protein tyrosine phosphatase function and the activation of ERBB receptors and downstream signaling pathways. Radiation-induced activation of extracellular regulated kinase (ERK)1/2 promoted the cleavage and release of paracrine ligands in carcinoma cells which caused re-activation of ERBB family receptors and intracellular signaling pathways. Blocking ERBB receptor phosphorylation or ERK1/2 pathway activity using small-molecule inhibitors of kinases for a short period of time following exposure (3 h) surprisingly protected tumor cells from the toxic effects of ionizing radiation. Prolonged exposure (48-72 h) of tumor cells to inhibition of ERBB receptor/ERK1/2 function enhanced radiosensitivity. In addition to ERBB receptor signaling, expression of activated forms of RAS family members and alterations in p53 mutational status are known to regulate radiosensitivity apparently independent of ERBB receptor function; however, changes in RAS or p53 mutational status, in isogenic HCT116 cells, were also noted to modulate the expression of ERBB receptors and ERBB receptor paracrine ligands. These alterations in receptor and ligand expression correlated with changes in the ability of HCT116 cells to activate ERK1/2 and AKT after irradiation, and to survive radiation exposure. Collectively, our data in multiple human carcinoma cell lines argues that tumor cells are dynamic and rapidly adapt to any single therapeutic challenge, for example, radiation and/or genetic manipulation e.g. loss of activated RAS function, to maintain tumor cell growth and viability.

Restoration of receptor-type protein tyrosine phosphatase eta function inhibits human pancreatic carcinoma cell growth in vitro and in vivo.

DEP-1/HPTPeta, a receptor-type protein tyrosine phosphatase, is a candidate tumor suppressor gene because its expression was blocked in rat and human thyroid transformed cells, and its restoration reverted their neoplastic phenotype. In addition, loss of DEP-1/HPTPeta heterozygosity has been described in mammary, lung and colon primary tumors. We now show that DEP-1/HPTPeta is drastically reduced in several cell lines originating from human epithelial pancreatic carcinomas compared with normal pancreatic tissue. We also show that the infection of AsPC1 and PSN1 cells with a recombinant adenovirus carrying r-PTPeta cDNA (the rat homolog of DEP-1/HPTPeta) inhibits their proliferation. Flow cytometric analysis of the infected cells demonstrated that restoration of r-PTPeta activity disrupts their cell cycle and leads to apoptosis. Finally, the growth of PSN1 xenograft tumors was blocked by the intratumoral injection of a recombinant adeno-associated virus carrying r-PTPeta. The data suggest that restoration of DEP-1/HPTPeta expression could be a useful tool for the gene therapy of human pancreatic cancers.

Emerging issues in receptor protein tyrosine phosphatase function: lifting fog or simply shifting?

Transmembrane (receptor) tyrosine phosphatases are intimately involved in responses to cell-cell and cell-matrix contact. Several important issues regarding the targets and regulation of this protein family are now emerging. For example, these phosphatases exhibit complex interactions with signaling pathways involving SRC family kinases, which result from their ability to control phosphorylation of both activating and inhibitory sites in these kinases and possibly also their substrates. Similarly, integrin signaling illustrates how phosphorylation of a single protein, or the activity of a pathway, can be controlled by multiple tyrosine phosphatases, attesting to the intricate integration of these enzymes in cellular regulation. Lastly, we are starting to appreciate the roles of intracellular topology, tyrosine phosphorylation and oligomerization among the many mechanisms regulating tyrosine phosphatase activity.

Molecular mechanism for the Shp-2 tyrosine phosphatase function in promoting growth factor stimulation of Erk activity.

We have previously shown that activation of extracellular signal-regulated kinase (Erk) by epidermal growth factor (EGF) treatment was significantly decreased in mouse fibroblast cells expressing a mutant Shp-2 molecule lacking 65 amino acids in the SH2-N domain, Shp-2(Delta46-110). To address the molecular mechanism for the positive role of Shp-2 in mediating Erk induction, we evaluated the activation of signaling components upstream of Erk in Shp-2 mutant cells. EGF-stimulated Ras, Raf, and Mek activation was significantly attenuated in Shp-2 mutant cells, suggesting that Shp-2 acts to promote Ras activation or to suppress the down-regulation of activated Ras. Biochemical analyses indicate that upon EGF stimulation, Shp-2 is recruited into a multiprotein complex assembled on the Gab1 docking molecule and that Shp-2 seems to exert its biological function by specifically dephosphorylating an unidentified molecule of 90 kDa in the complex. The mutant Shp-2(Delta46-110) molecule failed to participate in the Gab1-organized complex for dephosphorylation of p90, correlating with a defective activation of the Ras-Raf-Mek-Erk cascade in EGF-treated Shp-2 mutant cells. Evidence is also presented that Shp-2 does not appear to modulate the signal relay from EGF receptor to Ras through the Shc, Grb2, and Sos proteins. These results begin to elucidate the mechanism of Shp-2 function downstream of a receptor tyrosine kinase to promote the activation of the Ras-Erk pathway, with potential therapeutic applications in cancer treatment.

Receptor tyrosine phosphatases: the worm clears the picture.

Recent work on the Caenorhabditis elegans clr-1 gene shows that the receptor tyrosine phosphatase that it encodes negatively regulates a receptor tyrosine kinase related to mammalian fibroblast growth factor receptors. This opens up a promising system for investigating receptor tyrosine phosphatase function.

Dimerization-induced inhibition of receptor protein tyrosine phosphatase function through an inhibitory wedge.

The function and regulation of the receptorlike transmembrane protein tyrosine phosphatases (RPTPs) are not well understood. Ligand-induced dimerization inhibited the function of the epidermal growth factor receptor (EGFR)-RPTP CD45 chimera (EGFR-CD45) in T cell signal transduction. Properties of mutated EGFR-CD45 chimeras supported a general model for the regulation of RPTPs, derived from the crystal structure of the RPTPalpha membrane-proximal phosphatase domain. The phosphatase domain apparently forms a symmetrical dimer in which the catalytic site of one molecule is blocked by specific contacts with a wedge from the other.