scholarly journals Annexin A2 supports pulmonary microvascular integrity by linking vascular endothelial cadherin and protein tyrosine phosphatases

2017 ◽  
Vol 214 (9) ◽  
pp. 2535-2545 ◽  
Author(s):  
Min Luo ◽  
Elle C. Flood ◽  
Dena Almeida ◽  
LunBiao Yan ◽  
David A. Berlin ◽  
...  
Keyword(s):  
Pulmonary Edema ◽  
Tyrosine Phosphatase ◽  
Protein Tyrosine Phosphatases ◽  
Annexin A2 ◽  
Vascular Endothelial ◽  
Vascular Integrity ◽  
Protein Tyrosine ◽  
Vascular Endothelial Cadherin ◽  
Junction Protein ◽  
Alveolar Hypoxia

Relative or absolute hypoxia activates signaling pathways that alter gene expression and stabilize the pulmonary microvasculature. Alveolar hypoxia occurs in disorders ranging from altitude sickness to airway obstruction, apnea, and atelectasis. Here, we report that the phospholipid-binding protein, annexin A2 (ANXA2) functions to maintain vascular integrity in the face of alveolar hypoxia. We demonstrate that microvascular endothelial cells (ECs) from Anxa2−/− mice display reduced barrier function and excessive Src-related tyrosine phosphorylation of the adherens junction protein vascular endothelial cadherin (VEC). Moreover, unlike Anxa2+/+ controls, Anxa2−/− mice develop pulmonary edema and neutrophil infiltration in the lung parenchyma in response to subacute alveolar hypoxia. Mice deficient in the ANXA2-binding partner, S100A10, failed to demonstrate hypoxia-induced pulmonary edema under the same conditions. Further analyses reveal that ANXA2 forms a complex with VEC and its phosphatases, EC-specific protein tyrosine phosphatase (VE-PTP) and Src homology phosphatase 2 (SHP2), both of which are implicated in vascular integrity. In the absence of ANXA2, VEC is hyperphosphorylated at tyrosine 731 in response to vascular endothelial growth factor, which likely contributes to hypoxia-induced extravasation of fluid and leukocytes. We conclude that ANXA2 contributes to pulmonary microvascular integrity by enabling VEC-related phosphatase activity, thereby preventing vascular leak during alveolar hypoxia.

scholarly journals Vascular Endothelial Receptor Tyrosine Phosphatase: Identification of Novel Substrates Related to Junctions and a Ternary Complex with EPHB4 and TIE2

2019 ◽  
Vol 18 (10) ◽  
pp. 2058-2077 ◽  
Author(s):  
Hannes C. A. Drexler ◽  
Matthias Vockel ◽  
Christian Polaschegg ◽  
Maike Frye ◽  
Kevin Peters ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Tyrosine Kinase ◽  
Ternary Complex ◽  
Tyrosine Phosphatase ◽  
Cell Junctions ◽  
Receptor Protein ◽  
Spectrometric Analysis ◽  
Vascular Endothelial ◽  
Protein Tyrosine ◽  
Junction Protein

Vascular endothelial protein tyrosine phosphatase (VE-PTP, PTPRB) is a receptor type phosphatase that is crucial for the regulation of endothelial junctions and blood vessel development. We and others have shown recently that VE-PTP regulates vascular integrity by dephosphorylating substrates that are key players in endothelial junction stability, such as the angiopoietin receptor TIE2, the endothelial adherens junction protein VE-cadherin and the vascular endothelial growth factor receptor VEGFR2. Here, we have systematically searched for novel substrates of VE-PTP in endothelial cells by utilizing two approaches. First, we studied changes in the endothelial phosphoproteome on exposing cells to a highly VE-PTP-specific phosphatase inhibitor followed by affinity isolation and mass-spectrometric analysis of phosphorylated proteins by phosphotyrosine-specific antibodies. Second, we used a substrate trapping mutant of VE-PTP to pull down phosphorylated substrates in combination with SILAC-based quantitative mass spectrometry measurements. We identified a set of substrate candidates of VE-PTP, of which a remarkably large fraction (29%) is related to cell junctions. Several of those were found in both screens and displayed very high connectivity in predicted functional interaction networks. The receptor protein tyrosine kinase EPHB4 was the most prominently phosphorylated protein on VE-PTP inhibition among those VE-PTP targets that were identified by both proteomic approaches. Further analysis revealed that EPHB4 forms a ternary complex with VE-PTP and TIE2 in endothelial cells. VE-PTP controls the phosphorylation of each of these two tyrosine kinase receptors. Despite their simultaneous presence in a ternary complex, stimulating each of the receptors with their own specific ligand did not cross-activate the respective partner receptor. Our systematic approach has led to the identification of novel substrates of VE-PTP, of which many are relevant for the control of cellular junctions further promoting the importance of VE-PTP as a key player of junctional signaling.

scholarly journals Antagonism of VEGF-A–induced increase in vascular permeability by an integrin α3β1-Shp-1-cAMP/PKA pathway

Blood ◽  
2012 ◽  
Vol 120 (24) ◽  
pp. 4892-4902 ◽  
Author(s):  
Soo Hyeon Kim ◽  
Young-Rak Cho ◽  
Hyeon-Ju Kim ◽  
Joa Sub Oh ◽  
Eun-Kyung Ahn ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Vascular Permeability ◽  
Protein Tyrosine Phosphatase ◽  
Tyrosine Phosphatase ◽  
Vascular Endothelial ◽  
Protein Tyrosine ◽  
Vascular Endothelial Cadherin ◽  
Integrin Α3β1 ◽  
Kinase A

Abstract In cancer, VEGF-induced increase in vascular permeability results in increased interstitial pressure, reducing perfusion and increasing hypoxia, which reduce delivery of chemotherapeutic agents and increase resistance to ionizing radiation. Here, we show that both TIMP-2 and Ala + TIMP-2, a TIMP-2 mutant without matrix metalloproteinase inhibitory activity, antagonize the VEGF-A–induced increase in vascular permeability, both in vitro and in vivo. Like other agents known to preserve endothelial barrier function, TIMP-2 elevates cytosolic levels of cAMP and increases cytoskeletal-associated vascular endothelial cadherin in human microvascular endothelial cells. All of these effects are completely ablated by selective knockdown of integrin α3β1 expression, expression of a dominant negative protein tyrosine phosphatase Shp-1 mutant, administration of the protein tyrosine phosphatase inhibitor orthovanadate, or the adenylate cyclase inhibitor SQ22536. This TIMP-2–mediated inhibition of vascular permeability involves an integrin α3β1-Shp-1-cAMP/protein kinase A-dependent vascular endothelial cadherin cytoskeletal association, as evidenced by using siRNAs to integrin α3β1 and Shp-1, or treatment with Shp-1 inhibitor NSC87877 and protein kinase A inhibitor H89. Our results demonstrate the potential utility for TIMP-2 in cancer therapy through “normalization” of vascular permeability in addition to previously described antiangiogenic effects.

Design and Biological Evaluation of Novel Imidazolyl Flavonoids as Potent and Selective Protein Tyrosine Phosphatase Inhibitors

2020 ◽  
Vol 16 (4) ◽  
pp. 563-574 ◽  
Author(s):  
Rong Y. Han ◽  
Yu Ge ◽  
Ling Zhang ◽  
Qing M. Wang
Keyword(s):  
Protein Tyrosine Phosphatase ◽  
Tyrosine Phosphatase ◽  
Protein Tyrosine Phosphatases ◽  
Structural Features ◽  
Biological Evaluation ◽  
Cell Protein ◽  
Phosphatase Inhibitors ◽  
Protein Tyrosine ◽  
Therapeutic Development ◽  
Chemical Studies

Background: Protein tyrosine phosphatases 1B are considered to be a desirable validated target for therapeutic development of type II diabetes and obesity. Methods: A new series of imidazolyl flavonoids as potential protein tyrosine phosphatase inhibitors were synthesized and evaluated. Results: Bioactive results indicated that some synthesized compounds exhibited potent protein phosphatase 1B (PTP1B) inhibitory activities at the micromolar range. Especially, compound 8b showed the best inhibitory activity (IC50=1.0 µM) with 15-fold selectivity for PTP1B over the closely related T-cell protein tyrosine phosphatase (TCPTP). Cell viability assays indicated that 8b is cell permeable with lower cytotoxicity. Molecular modeling and dynamics studies revealed the reason for selectivity of PTP1B over TCPTP. Quantum chemical studies were carried out on these compounds to understand the structural features essential for activity. Conclusion: Compound 8b should be a potential selective PTP1B inhibitor.

scholarly journals Development of a Robust High-Throughput Screening Platform for Inhibitors of the Striatal-Enriched Tyrosine Phosphatase (STEP)

2021 ◽  
Vol 22 (9) ◽  
pp. 4417
Author(s):  
Lester J Lambert ◽  
Stefan Grotegut ◽  
Maria Celeridad ◽  
Palak Gosalia ◽  
Laurent JS De Backer ◽  
...  
Keyword(s):  
Drug Discovery ◽  
High Throughput ◽  
Tyrosine Kinases ◽  
High Throughput Screening ◽  
Kinase Inhibitors ◽  
Tyrosine Phosphatase ◽  
Protein Tyrosine Phosphatases ◽  
Synaptic Function ◽  
Label Free ◽  
Protein Tyrosine

Many human diseases are the result of abnormal expression or activation of protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs). Not surprisingly, more than 30 tyrosine kinase inhibitors (TKIs) are currently in clinical use and provide unique treatment options for many patients. PTPs on the other hand have long been regarded as “undruggable” and only recently have gained increased attention in drug discovery. Striatal-enriched tyrosine phosphatase (STEP) is a neuron-specific PTP that is overactive in Alzheimer’s disease (AD) and other neurodegenerative and neuropsychiatric disorders, including Parkinson’s disease, schizophrenia, and fragile X syndrome. An emergent model suggests that the increase in STEP activity interferes with synaptic function and contributes to the characteristic cognitive and behavioral deficits present in these diseases. Prior efforts to generate STEP inhibitors with properties that warrant clinical development have largely failed. To identify novel STEP inhibitor scaffolds, we developed a biophysical, label-free high-throughput screening (HTS) platform based on the protein thermal shift (PTS) technology. In contrast to conventional HTS using STEP enzymatic assays, we found the PTS platform highly robust and capable of identifying true hits with confirmed STEP inhibitory activity and selectivity. This new platform promises to greatly advance STEP drug discovery and should be applicable to other PTP targets.

scholarly journals Coordinated Regulation of Insulin Signaling by the Protein Tyrosine Phosphatases PTP1B and TCPTP

2005 ◽  
Vol 25 (2) ◽  
pp. 819-829 ◽  
Author(s):  
Sandra Galic ◽  
Christine Hauser ◽  
Barbara B. Kahn ◽  
Fawaz G. Haj ◽  
Benjamin G. Neel ◽  
...  
Keyword(s):  
Insulin Signaling ◽  
Tyrosine Phosphatase ◽  
Protein Tyrosine Phosphatases ◽  
Negative Regulator ◽  
Mitogen Activated Protein Kinase ◽  
Protein Tyrosine ◽  
Akt Activation ◽  
Protein Levels ◽  
Mitogen Activated Protein

ABSTRACT The protein tyrosine phosphatase PTP1B is a negative regulator of insulin signaling and a therapeutic target for type 2 diabetes. Our previous studies have shown that the closely related tyrosine phosphatase TCPTP might also contribute to the regulation of insulin receptor (IR) signaling in vivo (S. Galic, M. Klingler-Hoffmann, M. T. Fodero-Tavoletti, M. A. Puryer, T. C. Meng, N. K. Tonks, and T. Tiganis, Mol. Cell. Biol. 23:2096-2108, 2003). Here we show that PTP1B and TCPTP function in a coordinated and temporally distinct manner to achieve an overall regulation of IR phosphorylation and signaling. Whereas insulin-induced phosphatidylinositol 3-kinase/Akt signaling was prolonged in both TCPTP−/− and PTP1B−/− immortalized mouse embryo fibroblasts (MEFs), mitogen-activated protein kinase ERK1/2 signaling was elevated only in PTP1B-null MEFs. By using phosphorylation-specific antibodies, we demonstrate that both IR β-subunit Y1162/Y1163 and Y972 phosphorylation are elevated in PTP1B−/− MEFs, whereas Y972 phosphorylation was elevated and Y1162/Y1163 phosphorylation was sustained in TCPTP−/− MEFs, indicating that PTP1B and TCPTP differentially contribute to the regulation of IR phosphorylation and signaling. Consistent with this, suppression of TCPTP protein levels by RNA interference in PTP1B−/− MEFs resulted in no change in ERK1/2 signaling but caused prolonged Akt activation and Y1162/Y1163 phosphorylation. These results demonstrate that PTP1B and TCPTP are not redundant in insulin signaling and that they act to control both common as well as distinct insulin signaling pathways in the same cell.

scholarly journals In Vitro Characterization of the Bacillus subtilis Protein Tyrosine Phosphatase YwqE

2005 ◽  
Vol 187 (10) ◽  
pp. 3384-3390 ◽  
Author(s):  
Ivan Mijakovic ◽  
Lucia Musumeci ◽  
Lutz Tautz ◽  
Dina Petranovic ◽  
Robert A. Edwards ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Tyrosine Phosphatase ◽  
Protein Tyrosine Phosphatases ◽  
Class Ii ◽  
Gram Positive ◽  
Protein Tyrosine ◽  
Gram Positive Bacteria ◽  
New Family ◽  
In Vitro Characterization

ABSTRACT Both gram-negative and gram-positive bacteria possess protein tyrosine phosphatases (PTPs) with a catalytic Cys residue. In addition, many gram-positive bacteria have acquired a new family of PTPs, whose first characterized member was CpsB from Streptococcus pneumoniae. Bacillus subtilis contains one such CpsB-like PTP, YwqE, in addition to two class II Cys-based PTPs, YwlE and YfkJ. The substrates for both YwlE and YfkJ are presently unknown, while YwqE was shown to dephosphorylate two phosphotyrosine-containing proteins implicated in UDP-glucuronate biosynthesis, YwqD and YwqF. In this study, we characterize YwqE, compare the activities of the three B. subtilis PTPs (YwqE, YwlE, and YfkJ), and demonstrate that the two B. subtilis class II PTPs do not dephosphorylate the physiological substrates of YwqE.

Streptococcus sanguis-induced platelet activation involves two waves of tyrosine phosphorylation mediated by FcγRIIA and αIIbβ3

2005 ◽  
Vol 93 (05) ◽  
pp. 932-939 ◽  
Author(s):  
Caroline Pampolina ◽  
Archibald McNicol
Keyword(s):  
Signal Transduction ◽  
Platelet Aggregation ◽  
Tyrosine Phosphorylation ◽  
Platelet Activation ◽  
Tyrosine Phosphatase ◽  
Protein Tyrosine Phosphatases ◽  
Lag Phase ◽  
Dependent Manner ◽  
Protein Tyrosine ◽  
Streptococcus Sanguis

SummaryThe low-affinity IgG receptor, FcγRIIA, has been implicated in Streptococcus sanguis-induced platelet aggregation. Therefore, it is likely that signal transduction is at least partly mediated by FcγRIIA activation and a tyrosine kinase-dependent pathway. In this study the signal transduction mechanisms associated with platelet activation in response to the oral bacterium, S. sanguis were characterised. In the presence of IgG, S. sanguis strain 2017–78 caused the tyrosine phosphorylation of FcγRIIA 30s following stimulation, which led to the phosphorylation of Syk, LAT, and PLCγ2. These early events were dependent on Src family kinases but independent of either TxA2 or the engagement of the αIIbβ3 integrin. During the lag phase prior to platelet aggregation, FcγRIIA, Syk, LAT, and PLCγ2 were each dephosphorylated, but were re-phosphorylated as aggregation occurred. Platelet stimulation by 2017–78 also induced the tyrosine phosphorylation of PECAM-1, an ITIM-containing receptor that recruits protein tyrosine phosphatases. PECAM-1 co-precipitated with the protein tyrosine phosphatase SHP-1 in the lag phase. SHP-1 was also maximally tyrosine phosphorylated during this phase, suggesting a possible role for SHP-1 in the observed dephosphorylation events. As aggregation occurred, SHP-1 was dephosphorylated, while FcγRIIA, Syk, LAT, and PLCγ2 were rephosphorylated in an RGDS-sensitive, and therefore αIIbβ3-dependent, manner. Additionally, TxA2 release, 5-hydro-xytryptamine secretion and phosphatidic acid formation were all blocked by RGDS. Aspirin also abolished these events, but only partially inhibited αIIbβ3-mediated re-phosphorylation. Therefore, S.sanguis-bound IgG cross links FcγRIIA and initiates a signaling pathway that is down-regulated by PECAM-1-bound SHP-1. Subsequent engagement of αIIbβ3 leads to SHP-1 dephosphorylation permiting a second wave of signaling leading to TxA2 release and consequent platelet aggregation.

scholarly journals Gata2 is required for HSC generation and survival

2013 ◽  
Vol 210 (13) ◽  
pp. 2843-2850 ◽  
Author(s):  
Emma de Pater ◽  
Polynikis Kaimakis ◽  
Chris S. Vink ◽  
Tomomasa Yokomizo ◽  
Tomoko Yamada-Inagawa ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Transcription Factor ◽  
Developmental Stages ◽  
Vascular Endothelial ◽  
Vascular Integrity ◽  
Hematopoietic Stem ◽  
Vascular Endothelial Cadherin ◽  
Umbilical Arteries ◽  
Specific Deletion

Knowledge of the key transcription factors that drive hematopoietic stem cell (HSC) generation is of particular importance for current hematopoietic regenerative approaches and reprogramming strategies. Whereas GATA2 has long been implicated as a hematopoietic transcription factor and its dysregulated expression is associated with human immunodeficiency syndromes and vascular integrity, it is as yet unknown how GATA2 functions in the generation of HSCs. HSCs are generated from endothelial cells of the major embryonic vasculature (aorta, vitelline, and umbilical arteries) and are found in intra-aortic hematopoietic clusters. In this study, we find that GATA2 function is essential for the generation of HSCs during the stage of endothelial-to-hematopoietic cell transition. Specific deletion of Gata2 in Vec (Vascular Endothelial Cadherin)-expressing endothelial cells results in a deficiency of long-term repopulating HSCs and intra-aortic cluster cells. By specific deletion of Gata2 in Vav-expressing hematopoietic cells (after HSC generation), we further show that GATA2 is essential for HSC survival. This is in contrast to the known activity of the RUNX1 transcription factor, which functions only in the generation of HSCs, and highlights the unique requirement for GATA2 function in HSCs throughout all developmental stages.

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