Expression Of Protein Tyrosine Phosphatase Research Articles

Protein Tyrosine Phosphatase Expression Profile of Rheumatoid Arthritis Fibroblast‐like Synoviocytes: A Novel Role of SH2 Domain–Containing Phosphatase 2 as a Modulator of Invasion and Survival

The fibroblast-like synoviocytes (FLS) in the synovial intimal lining of the joint are key mediators of inflammation and joint destruction in rheumatoid arthritis (RA). In RA, these cells aggressively invade the extracellular matrix, producing cartilage-degrading proteases and inflammatory cytokines. The behavior of FLS is controlled by multiple interconnected signal transduction pathways involving reversible phosphorylation of proteins on tyrosine residues. However, little is known about the role of the protein tyrosine phosphatases (PTPs) in FLS function. This study was undertaken to explore the expression of all of the PTP genes (the PTPome) in FLS. A comparative screening of the expression of the PTPome in FLS from patients with RA and patients with osteoarthritis (OA) was conducted. The functional effect on RA FLS of SH2 domain-containing phosphatase 2 (SHP-2), a PTP that was up-regulated in RA, was then analyzed by knockdown using cell-permeable antisense oligonucleotides. PTPN11 was overexpressed in RA FLS compared to OA FLS. Knockdown of PTPN11, which encodes SHP-2, reduced the invasion, migration, adhesion, spreading, and survival of RA FLS. Additionally, signaling in response to growth factors and inflammatory cytokines was impaired by SHP-2 knockdown. RA FLS that were deficient in SHP-2 exhibited decreased activation of focal adhesion kinase and mitogen-activated protein kinases. These findings indicate that SHP-2 has a novel role in mediating human FLS function and suggest that it promotes the invasiveness and survival of RA FLS. Further investigation may reveal SHP-2 to be a candidate therapeutic target for RA.

Feeding a DHA-enriched diet increases skeletal muscle protein synthesis in growing pigs: association with increased skeletal muscle insulin action and local mRNA expression of insulin-like growth factor 1

Dietary n-3 PUFA have been demonstrated to promote muscle growth in growing animals. In the present study, fractional protein synthesis rates (FSR) in the skeletal muscle of growing pigs fed a DHA-enriched (DE) diet (DE treatment) or a soyabean oil (SO) diet (SO treatment) were evaluated in the fed and feed-deprived states. Feeding-induced increases in muscle FSR, as well as the activation of the mammalian target of rapamycin and protein kinase B, were higher in the DE treatment as indicated by the positive interaction between diet and feeding. In the fed state, the activation of eIF4E-binding protein 1 in the skeletal muscle of pigs on the DE diet was higher than that in pigs on the SO diet (P<0·05). Feeding the DE diet increased muscle insulin-like growth factor 1 (IGF-1) expression (P<0·05) and insulin action (as demonstrated by increased insulin receptor (IR) phosphorylation, P<0·05), resulting in increased IR substrate 1 activation in the fed state. However, no difference in plasma IGF-1 concentration or hepatic IGF-1 expression between the two treatments was associated. The increased IGF-1 expression in the DE treatment was associated with increased mRNA expression of the signal transducer and activator of transcription 5A and decreased mRNA expression of protein tyrosine phosphatase, non-receptor type 3 in skeletal muscle. Moreover, mRNA expression of protein tyrosine phosphatase, non-receptor type 1 (PTPN1), the activation of PTPN1 and the activation of NF-κB in muscle were significantly lower in the DE treatment (P<0·05). The results of the present study suggest that feeding a DE diet increased feeding-induced muscle protein synthesis in growing pigs, and muscle IGF-1 expression and insulin action were involved in this action.

Expression of protein-tyrosine phosphatases in Acute Myeloid Leukemia cells: FLT3 ITD sustains high levels of DUSP6 expression

Protein-tyrosine phosphatases (PTPs) are important regulators of cellular signaling and changes in PTP activity can contribute to cell transformation. Little is known about the role of PTPs in Acute Myeloid Leukemia (AML). The aim of this study was therefore to establish a PTP expression profile in AML cells and to explore the possible role of FLT3 ITD (Fms-like tyrosine kinase 3 with internal tandem duplication), an important oncoprotein in AML for PTP gene expression. PTP mRNA expression was analyzed in AML cells from patients and in cell lines using a RT-qPCR platform for detection of transcripts of 92 PTP genes. PTP mRNA expression was also analyzed based on a public microarray data set for AML patients. Highly expressed PTPs in AML belong to all PTP subfamilies. Very abundantly expressed PTP genes include PTPRC, PTPN2, PTPN6, PTPN22, DUSP1, DUSP6, DUSP10, PTP4A1, PTP4A2, PTEN, and ACP1. PTP expression was further correlated with the presence of FLT3 ITD, focusing on a set of highly expressed dual-specificity phosphatases (DUSPs). Elevated expression of DUSP6 in patients harboring FLT3 ITD was detected in this analysis. The mechanism and functional role of FLT3 ITD-mediated upregulation of DUSP6 was then explored using pharmacological inhibitors of FLT3 ITD signal transduction and si/shRNA technology in human and murine cell lines. High DUSP6 expression was causally associated with the presence of FLT3 ITD and dependent on FLT3 ITD kinase activity and ERK signaling. DUSP6 depletion moderately increased ERK1/2 activity but attenuated FLT3 ITD-dependent cell proliferation of 32D cells. In conclusion, DUSP6 may play a contributing role to FLT3 ITD-mediated cell transformation.

Regulation of signaling molecules associated with insulin action, insulin secretion and pancreatic β-cell mass in the hypoglycemic effects of Korean red ginseng in Goto-Kakizaki rats

Ethnopharmacological relevanceKorean red ginseng (KRG) has long history as herbal remedy for antidiabetic effect. Aim of the studyTo study molecular mechanisms by which KRG ameliorates diabetes mellitus, we investigated whether the supplementation with the aqueous extract of KRG as a dietary admixture (1%, w/w) regulates the expressions of signaling molecules that are associated with insulin action, insulin secretion and pancreatic β-cell mass in spontaneously diabetic Goto-Kakizaki (GK) rats. MethodsAn aqueous extract of KRG was supplemented for the estimated dosage to be 0.2g/kg rat/day beginning at 5 weeks of age for 12 weeks in male GK rats. Plasma glucose levels were measured every 4 weeks. The expressions of signaling molecules that are associated with insulin action, insulin secretion and β-cell mass in tissues were determined by Western blotting. ResultsThe 12-week supplementation with KRG significantly (P<0.05) decreased blood glucose compared to control. It up-regulated the expression of glucose transporter (GLUT) 4 in adipose tissue, and down-regulated the expression of protein tyrosine phosphatases (PTP)-1B in adipose tissue and skeletal muscle. It also up-regulated the expression of insulin and down-regulated the expression of uncoupling protein (UCP) 2, Bax and poly (ADP-ribose) polymerase (PARP) in pancreas. ConclusionsThese results suggest that GLUT4, PTP-1B, insulin, UCP2, Bax and PARP may be the primary targets of KRG that result in increase in insulin action and in insulin secretion, and decrease in β-cell mass, and that cause the normalization in glucose homeostasis.

Icariside II Induces Apoptosis in U937 Acute Myeloid Leukemia Cells: Role of Inactivation of STAT3-Related Signaling

BackgroundThe aim of this study is to determine anti-cancer effect of Icariside II purified from the root of Epimedium koreanum Nakai on human acute myeloid leukemia (AML) cell line U937.Methodology/Principal FindingsIcariside II blocked the growth U937 cells in a dose- and time-dependent manner. In this anti-proliferation process, this herb compound rendered the cells susceptible to apoptosis, manifested by enhanced accumulation of sub-G1 cell population and increased the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive cells. Icariside II was able to activate caspase-3 and cleaved poly (ADP-ribose) polymerase (PARP) in a time-dependent manner. Concurrently, the anti-apoptotic proteins, such as bcl-xL and survivin in U937 cells, were downregulated by Icariside II. In addition, Icariside II could inhibit STAT3 phosphorylation and function and subsequently suppress the activation of Janus activated kinase 2 (JAK2), the upstream activators of STAT3, in a dose- and time-dependent manner. Icariside II also enhanced the expression of protein tyrosine phosphatase (PTP) SH2 domain-containing phosphatase (SHP)-1, and the addition of sodium pervanadate (a PTP inhibitor) prevented Icariside II-induced apoptosis as well as STAT3 inactivation in STAT3 positive U937 cells. Furthermore, silencing SHP-1 using its specific siRNA significantly blocked STAT3 inactivation and apoptosis induced by Icariside II in U937 cells.Conclusions/SignificanceOur results demonstrated that via targeting STAT3-related signaling, Icariside II sensitizes U937 cells to apoptosis and perhaps serves as a potent chemotherapeutic agent for AML.

SHP2 Converts Parafibromin/Cdc73 from Tumor Suppressor to Oncoprotein

SHP2 is a ubiquitously expressed protein tyrosine phosphatase encoded by the PTPN11 gene. Mutations in PTPN11 are associated with the hereditary developmental disorder Noonan syndrome in addition to adult and juvenile leukemia. SHP2 plays a central role in cell signaling events that promote growth and motility and is required for activation of the RAS signaling pathway. Deregulation of SHP also has been implicated in the development of multiple human malignancies. In a recent article, Takahashi and colleagues identified a unique role for SHP2 in the nucleus, as the determinant of the function of parafibromin/Cdc73, a component of the nuclear RNA polymerase II-associated factor (PAF) complex. Parafibromin has opposing functions that are context dependent. When complexed with the histone methyltransferase SUV39H1, parafibromin acts as a tumor suppressor that inhibits the transcription of cyclin D1 and c-myc. However, upon binding β-catenin, parafibromin switches to an oncogene that activates the promitogenic Wnt signaling pathway. The authors discovered that parafibromin is a unique substrate of SHP2 and that dephosphorylation by SHP2 determines its binding partner and therefore its nuclear function. In the absence of SHP2, parafibromin was tyrosine-phosphorylated and complexed with SUV39H1. Upon tyrosine dephosphorylation by SHP2, parafibromin preferentially bound to and trapped β-catenin in the nucleus. The formation and nuclear accumulation of the stable parafibromin/β-catenin complex was able to override parafibromin/SUV39H1-mediated repression and led to the induction of Wnt target genes. Overall, the important finding that SHP2 activity determines whether parafibromin behaves as a tumor suppressor or an oncogene offers mechanistic insight into how the deregulation of SHP2 may play a causative role in the development of neoplasms and developmental malformations.Takahashi A, Tsutsumi R, Kikuchi I, Obuse C, Saito Y, Seidi A, et al. SHP2 tyrosine phosphatase converts parafibromin/Cdc73 from a tumor suppressor to an oncogenic driver. Mol Cell 2011;43:45–56.Note: Research Watch is written by Cancer Discovery Editors. Readers are encouraged to consult the articles referred to in each item for full details on the findings described.

Novel protein tyrosine phosphatase 1B inhibitors: interaction requirements for improved intracellular efficacy in type 2 diabetes mellitus and obesity control

Resistance to the hormones insulin and leptin are hallmarks in common for type 2 diabetes mellitus and obesity. Both conditions are associated with increased activity and expression of protein tyrosine phosphatase (PTP)1B. Therefore, inhibition of PTP1B activity or down-regulation of its expression should ameliorate insulin and leptin resistance, and may hold therapeutic utility in type 2 diabetes mellitus and obesity control. This background has motivated the fervent search for PTP1B inhibitors, carried out in the recent years. The purpose of this review is to provide the most recent advances in understanding the structural details of PTP1B molecule relevant to the interactions with inhibitors, and the progress towards compounds with enhanced membrane permeability, affinity, specificity, and potency on intracellular PTP1B; several inhibitors of benefit in type 2 diabetes mellitus and obesity control are presented and discussed.

Protein Tyrosine Phosphatase SHP-2 (PTPN11) in Hematopoiesis and Leukemogenesis.

SHP-2 (PTPN11), a ubiquitously expressed protein tyrosine phosphatase, is critical for hematopoietic cell development and function owing to its essential role in growth factor/cytokine signaling. More importantly, germline and somatic mutations in this phosphatase are associated with Noonan syndrome, Leopard syndrome, and childhood hematologic malignancies. The molecular mechanisms by which SHP-2 mutations induce these diseases are not fully understood, as the biochemical bases of SHP-2 functions still remain elusive. Further understanding SHP-2 signaling activities and identification of its interacting proteins/substrates will shed light on the pathogenesis of PTPN11-associated hematologic malignancies, which, in turn, may lead to novel therapeutics for these diseases.

Striatal-Enriched Protein Tyrosine Phosphatase Expression and Activity in Huntington's Disease: A STEP in the Resistance to Excitotoxicity

Striatal-enriched protein tyrosine phosphatase (STEP) is highly expressed in striatal projection neurons, the neuronal population most affected in Huntington's disease. Here, we examined STEP expression and phosphorylation, which regulates its activity, in N-terminal exon-1 and full-length mutant huntingtin mouse models. R6/1 mice displayed reduced STEP protein levels in the striatum and cortex, whereas its phosphorylation was increased in the striatum, cortex, and hippocampus. The early increase in striatal STEP phosphorylation levels correlated with a deregulation of the protein kinase A pathway, and decreased calcineurin activity at later stages further contributes to an enhancement of STEP phosphorylation and inactivation. Accordingly, we detected an accumulation of phosphorylated ERK2 and p38, two targets of STEP, in R6/1 mice striatum at advanced stages of the disease. Activation of STEP participates in excitotoxic-induced cell death. Because Huntington's disease mouse models develop resistance to excitotoxicity, we analyzed whether decreased STEP activity was involved in this process. After intrastriatal quinolinic acid (QUIN) injection, we detected higher phosphorylated STEP levels in R6/1 than in wild-type mice, suggesting that STEP inactivation could mediate neuroprotection in R6/1 striatum. In agreement, intrastriatal injection of TAT-STEP increased QUIN-induced cell death. R6/2, Tet/HD94, and Hdh(Q7/Q111) mice striatum also displayed decreased STEP protein and increased phosphorylation levels. In Tet/HD94 mice striatum, mutant huntingtin transgene shutdown reestablished STEP expression. In conclusion, the STEP pathway is severely downregulated in the presence of mutant huntingtin and may participate in compensatory mechanisms activated by striatal neurons that lead to resistance to excitotoxicity.

Inhibition of JAK1/STAT3 signaling mediates compound K-induced apoptosis in human multiple myeloma U266 cells

Signal transducer and activator of transcription 3 (STAT3) is an oncogenic transcription factor implicated in carcinogenesis. Here, the role of STAT3 pathway in the antitumor activity of an active ginseng saponin metabolite compound K (CK) was investigated in human multiple myeloma U266 cells. CK increased the cytotoxicity, accumulated the sub-G1 DNA population, cleaved poly (ADP-ribose) polymerase (PARP) and activated caspase-3 in U266 cells. Interestingly, CK inhibited phosphorylation of STAT3 and its upstream activators, the Janus activated kinase 1 (JAK1), but not JAK2. Furthermore, CK enhanced the expression of protein tyrosine phosphatase (PTP) SHP-1, but not PTEN. Additionally, CK down-regulated STAT3 target genes bcl-xL, bcl-2, survivin, cyclin E and cyclin D1. Conversely, PTP inhibitor pervanadate reversed CK-mediated STAT3 inactivation and cleavages of caspase-3 and PARP. Overall, our findings demonstrate that JAK1/STAT3 signaling mediates CK-induced apoptosis in U266 cells and also suggest the chemopreventive potential of CK for treatment of multiple myeloma.

Somatostatin receptor-3 mediated intracellular signaling and apoptosis is regulated by its cytoplasmic terminal

In the present study, we describe the role of cytoplasmic terminal (C-tail) domain in regulating coupling to adenylyl cyclase, signaling, and apoptosis in human embryonic kidney (HEK-293) cells transfected with wild type (wt)-hSSTR3 and C-tail deleted mutants. Cells transfected with wt-hSSTR3 and C-tail mutants show comparable membrane expression; however, display decreased expression in presence of agonist. wt-hSSTR3 exists as preformed homodimer at cell surface in basal conditions and decreases in response to agonist. Cells expressing C-tail mutants also show evidence of homodimerization with the same intensity as wt-hSSTR3. The agonist-dependent inhibition of cyclic adenosine monophosphate (cAMP) was lost in cells expressing C-tail mutants. Agonist treatment in cells expressing wt-hSSTR3 resulted in inhibition of cell proliferation, increased expression of PARP-1, and TUNEL positivity in proliferating cell nuclear antigen (PCNA)-positive cells. The agonist mediated increase in membrane expression of protein tyrosine phosphatase (PTP) seen with wt-hSSTR3 was diminished in C-tail mutants, which was accompanied with the loss of receptor's ability to induce apoptosis. Taken together, our data provide new insights into C-tail-dependent regulation of cell signaling and apoptosis by hSSTR3.

Hypoxia Enhances Platelet-derived Growth Factor Signaling in the Pulmonary Vasculature by Down-Regulation of Protein Tyrosine Phosphatases

Platelet-derived growth factor (PDGF) plays a pivotal role in the pathobiology of pulmonary hypertension (PH) because it promotes pulmonary vascular remodeling. PH is frequently associated with pulmonary hypoxia. To investigate whether hypoxia alters PDGF β receptor (βPDGFR) signaling in the pulmonary vasculature. The impact of chronic hypoxia on signal transduction by the βPDGFR was measured in human pulmonary arterial smooth muscle cells (hPASMC) in vitro, and in mice with hypoxia-induced PH in vivo. Chronic hypoxia significantly enhanced PDGF-BB-dependent proliferation and chemotaxis of hPASMC. Pharmacologic inhibition of PI3 kinase (PI3K) and PLCγ abrogated these events under both normoxia and hypoxia. Although hypoxia did not affect βPDGFR expression, it increased the ligand-induced tyrosine phosphorylation of the receptor, particularly at binding sites for PI3K (Y751) and PLCγ (Y1021). The activated βPDGFR is dephosphorylated by protein tyrosine phosphatases (PTPs). Interestingly, hypoxia decreased expression of numerous PTPs (T cell PTP, density-enhanced phosphatase-1, PTP1B, and SH2 domain-containing phosphatase-2), resulting in reduced PTP activity. Hypoxia-inducible factor (HIF)-1α is involved in this regulation of gene expression, because hypoxia-induced βPDGFR hyperphosphorylation and PTP down-regulation were abolished by HIF-1α siRNA and by the HIF-1α inhibitor 2-methoxyestradiol. βPDGFR hyperphosphorylation and PTP down-regulation were also present in vivo in mice with chronic hypoxia-induced PH. Hypoxia reduces expression and activity of βPDGFR-antagonizing PTPs in a HIF-1α-dependent manner, thereby enhancing receptor activation and proliferation and chemotaxis of hPASMC. Because hyperphosphorylation of the βPDGFR and down-regulation of PTPs occur in vivo, this mechanism likely has significant impact on the development and progression of PH and other hypoxia-associated diseases.

Development of severe skeletal defects in induced SHP-2-deficient adult mice: a model of skeletal malformation in humans with SHP-2 mutations

SUMMARYSHP-2 (encoded by PTPN11) is a ubiquitously expressed protein tyrosine phosphatase required for signal transduction by multiple different cell surface receptors. Humans with germline SHP-2 mutations develop Noonan syndrome or LEOPARD syndrome, which are characterized by cardiovascular, neurological and skeletal abnormalities. To study how SHP-2 regulates tissue homeostasis in normal adults, we used a conditional SHP-2 mouse mutant in which loss of expression of SHP-2 was induced in multiple tissues in response to drug administration. Induced deletion of SHP-2 resulted in impaired hematopoiesis, weight loss and lethality. Most strikingly, induced SHP-2-deficient mice developed severe skeletal abnormalities, including kyphoses and scolioses of the spine. Skeletal malformations were associated with alterations in cartilage and a marked increase in trabecular bone mass. Osteoclasts were essentially absent from the bones of SHP-2-deficient mice, thus accounting for the osteopetrotic phenotype. Studies in vitro revealed that osteoclastogenesis that was stimulated by macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor kappa B ligand (RANKL) was defective in SHP-2-deficient mice. At least in part, this was explained by a requirement for SHP-2 in M-CSF-induced activation of the pro-survival protein kinase AKT in hematopoietic precursor cells. These findings illustrate an essential role for SHP-2 in skeletal growth and remodeling in adults, and reveal some of the cellular and molecular mechanisms involved. The model is predicted to be of further use in understanding how SHP-2 regulates skeletal morphogenesis, which could lead to the development of novel therapies for the treatment of skeletal malformations in human patients with SHP-2 mutations.

Identification and Expression of the Family of Classical Protein-Tyrosine Phosphatases in Zebrafish

Protein-tyrosine phosphatases (PTPs) have an important role in cell survival, differentiation, proliferation, migration and other cellular processes in conjunction with protein-tyrosine kinases. Still relatively little is known about the function of PTPs in vivo. We set out to systematically identify all classical PTPs in the zebrafish genome and characterize their expression patterns during zebrafish development. We identified 48 PTP genes in the zebrafish genome by BLASTing of human PTP sequences. We verified all in silico hits by sequencing and established the spatio-temporal expression patterns of all PTPs by in situ hybridization of zebrafish embryos at six distinct developmental stages. The zebrafish genome encodes 48 PTP genes. 14 human orthologs are duplicated in the zebrafish genome and 3 human orthologs were not identified. Based on sequence conservation, most zebrafish orthologues of human PTP genes were readily assigned. Interestingly, the duplicated form of ptpn23, a catalytically inactive PTP, has lost its PTP domain, indicating that PTP activity is not required for its function, or that ptpn23b has lost its PTP domain in the course of evolution. All 48 PTPs are expressed in zebrafish embryos. Most PTPs are maternally provided and are broadly expressed early on. PTP expression becomes progressively restricted during development. Interestingly, some duplicated genes retained their expression pattern, whereas expression of other duplicated genes was distinct or even mutually exclusive, suggesting that the function of the latter PTPs has diverged. In conclusion, we have identified all members of the family of classical PTPs in the zebrafish genome and established their expression patterns. This is the first time the expression patterns of all members of the large family of PTP genes have been established in a vertebrate. Our results provide the first step towards elucidation of the function of the family of classical PTPs.

Grape powder extract attenuates tumor necrosis factor α-mediated inflammation and insulin resistance in primary cultures of human adipocytes

Grapes are rich in phenolic phytochemicals that possess anti-oxidant and anti-inflammatory properties. However, the ability of grape powder extract (GPE) to prevent inflammation and insulin resistance in human adipocytes caused by tumor necrosis factor α (TNFα), a cytokine elevated in plasma and white adipose tissue (WAT) of obese, diabetic individuals, is unknown. Therefore, we examined the effects of GPE on markers of inflammation and insulin resistance in primary cultures of newly differentiated human adipocytes treated with TNFα. We found that GPE attenuated TNFα-induced expression of inflammatory genes including interleukin (IL)-6, IL-1β, IL-8, monocyte chemoattractant protein (MCP)-1, cyclooxygenase (COX)-2 and Toll-like receptor (TLR)-2. GPE attenuated TNFα-mediated activation of extracellular signal-related kinase (ERK) and c-Jun NH 2-terminal kinase (JNK) and activator protein-1 (AP-1, i.e., c-Jun). GPE also attenuated TNFα-mediated IκBα degradation and nuclear factor-kappa B (NF-κB) activity. Finally, GPE prevented TNFα-induced expression of protein tyrosine phosphatase (PTP)-1B and phosphorylation of serine residue 307 of insulin receptor substrate-1 (IRS-1), which are negative regulators of insulin sensitivity, and suppression of insulin-stimulated glucose uptake. Taken together, these data demonstrate that GPE attenuates TNFα-mediated inflammation and insulin resistance in human adipocytes, possibly by suppressing the activation of ERK, JNK, c-Jun and NF-κB.

Abstract 5774: Hypoxia Enhances Platelet-derived Growth Factor Signaling in the Pulmonary Vasculature by HIF-1 α -dependent Downregulation of Protein Tyrosine Phosphatases

Background: Platelet-derived growth factor (PDGF) plays a critical role in the development of pulmonary hypertension (PH) as it promotes pulmonary vascular remodeling. PH is frequently associated with severe pulmonary hypoxia. Here we investigated whether hypoxia alters PDGF β receptor ( β PDGFR) signaling. Methods and Results: Chronic hypoxia enhanced PDGF-BB-dependent proliferation and chemotaxis of human pulmonary arterial smooth muscle cells (hPASMC). Pharmacological inhibition of PI3 kinase (PI3K) and PLC γ abrogated these events both under normoxia and hypoxia. While hypoxia did not affect β PDGFR expression, it increased the ligand-induced tyrosine phosphorylation of the receptor, particularly at the binding sites for PI3K (Y751) and PLC γ (Y1021). The activated β PDGFR is dephosphorylated by protein tyrosine phosphatases (PTPs). Strikingly, hypoxia decreased the expression of numerous PTPs (TC-PTP, DEP-1, PTP1B, SHP-2), resulting in reduced PTP activity. Putative hypoxia-inducible factor (HIF)-1 α binding sites were identified in the promoter regions of the downregulated PTPs, and hypoxia-induced β PDGFR hyperphosphorylation and PTP downregulation were abolished by HIF-1 α siRNA and by the HIF-1 α inhibitor 2-methoxyestradiol. Immunohistochemistry and quantitative RT-PCR demonstrated that β PDGFR hyperphosphorylation and PTP downregulation were also present in vivo in mice with chronic hypoxia-induced PH. Conclusions: Hypoxia reduces the expression and activity of β PDGFR-antagonizing PTPs in a HIF-1 α -dependent manner, thereby enhancing receptor activation and proliferation/chemotaxis of hPASMC. As hyperphosphorylation of the β PDGFR and downregulation of PTPs also occur in vivo, this mechanism likely has significant impact on the development and progression of PH and other proliferative diseases.

Antibody binding to cell surface amyloid precursor protein induces neuronal injury by deregulating the phosphorylation of focal adhesion signaling related proteins

The biological function of full-length amyloid-β protein precursor (APP), the precursor of Aβ, is not fully understood. Mounting studies reported that antibody binding to cell surface APP causes neuronal injury. However, the mechanism of cell surface APP mediating neuronal injury remains to be determined. Colocalization of APP with integrin on cell surface leads us to suppose that focal adhesion (FA) related mechanism is involved in surface APP-mediated neuronal injury. In the present study, results demonstrated that primary cultured neurons treated with antibody against APP-N-terminal not only caused neuronal injury and aberrant morphologic changes of neurite, but also induced reaction of FA proteins appearing an acute increase then decrease pattern. Moreover, the elevation of tyrosine phosphorylation of FA proteins including paxillin and focal adhesion kinase (FAK), and down-regulated expression of protein tyrosine phosphatase (PTP1B) induced by APP antibody were prevented by inhibitor of Src protein kinases 4-amino-5-(4-chlorophenyl)-7(t-butyl) pyrazol (3,4-D) pyramide (PP2) and G protein inhibitor pertussis toxin (PTX), implying that Src family kinase and G protein play roles in APP-induced FA signals. In addition, pretreatment with PTX and PP2 was able to suppress APP-antibody induced neuronal injury. Taken together, the results suggest a novel mechanism for APP mediating neuronal injury through deregulating FA signals.

Protein tyrosine phosphatases expression during development of mouse superior colliculus

Protein tyrosine phosphatases (PTPs) are key regulators of different processes during development of the central nervous system. However, expression patterns and potential roles of PTPs in the developing superior colliculus remain poorly investigated. In this study, a degenerate primer-based reverse transcription-polymerase chain reaction (RT-PCR) approach was used to isolate seven different intracellular PTPs and nine different receptor-type PTPs (RPTPs) from embryonic E15 mouse superior colliculus. Subsequently, the expression patterns of 11 PTPs (TC-PTP, PTP1C, PTP1D, PTP-MEG2, PTP-PEST, RPTPJ, RPTPε, RPTPRR, RPTPσ, RPTPκ and RPTPγ) were further analyzed in detail in superior colliculus from embryonic E13 to postnatal P20 stages by quantitative real-time RT-PCR, Western blotting and immunohistochemistry. Each of the 11 PTPs exhibits distinct spatiotemporal regulation of mRNAs and proteins in the developing superior colliculus suggesting their versatile roles in genesis of neuronal and glial cells and retinocollicular topographic mapping. At E13, additional double-immunohistochemical analysis revealed the expression of PTPs in collicular nestin-positive neural progenitor cells and RC-2-immunoreactive radial glia cells, indicating the potential functional importance of PTPs in neurogenesis and gliogenesis.Electronic supplementary materialThe online version of this article (doi:10.1007/s00221-009-1963-6) contains supplementary material, which is available to authorized users.

Vascular PTPs: Current developments and challenges for exploitation in Type 2 diabetes-associated vascular dysfunction

Protein Tyrosine Phosphatases (PTPs) are important contributors to vascular cells normal function, by balancing signaling proteins activation exerted by phosphorylating kinases. Type 2 diabetes related insults, such as hyperglycemia, oxidative stress, and insulin resistance disturb the phosphorylation/dephosphorylation equilibrium towards an abnormal augmented phosphorylation of signaling proteins associated with changes in PTPs expression, enzymatic activity and interaction with cellular substrates. We briefly review here: (i) the new findings on receptor and non-receptor PTPs and their role in vascular cells, (ii) several data on oxidation and phosphorylation of these molecules in endothelial and smooth muscle cells, (iii) vascular PTPs intrinsic activity and dysregulation under the insults of diabetic milieu, and (iv) the potential use of PTPs and their inhibitors as therapeutic targets in Type 2 diabetes-associated vascular dysfunction.

"Shping 2" different cellular localizations - a potential new player in aging processes

The functions of the ubiquitously expressed protein tyrosine phosphatase Shp-2 are dependent on its localization. Cytosolic Shp-2 is known to modulate different pathways involved in cell growth, cell development, tissue inflammation and cellular chemotaxis. But Shp-2 is also localized in the nucleus and the mitochondria. Nuclear Shp-2 forms a complex with the signal transducer and activator of transcription 5 (STAT5) which then binds to DNA and regulates transcription of milk genes. In contrast, nuclear Shp-2 dephosphorylates STAT1 and thereby inhibits gene transcription. In addition, it counteracts the oxidative stress dependent nuclear export of Telomerase Reverse Transcriptase (TERT) mediated by members of the Src kinase family, a process leading to replicative senescence. For the recently found mitochondrial Shp-2 an involvement in the regulation of the cellular redox balance is discussed. Shp-2 shows the ability to regulate reactive oxygen species formation in the mitochondria. There are hints that mitochondrial Shp-2 and Src are involved in the regulation of respiratory chain activity. Since a substantial fraction of TERT has been found in the mitochondria, it is hypothesized that mitochondrial Shp-2 acts as a positive regulator of TERT in the mitochondria, similar to its nuclear role. Taken together, Shp-2 seems to be a new player in aging processes.