Deoxygenation and elevation of intracellular magnesium induce tyrosine phosphorylation of band 3 in human erythrocytes

Barbul, Alexander; Zipser, Yehudit; Nachles, Alexander; Korenstein, Rafi

doi:10.1016/s0014-5793(99)00822-4

Cited by 44 publications

(41 citation statements)

References 21 publications

(29 reference statements)

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“…In this study, we showed that the preexistence of Cys S-nitrosothiol prevented ROS-induced irreversible oxidation of PTP1B, which may be a mechanism for NO-mediated cytoprotective effect under pathological conditions, such as I/R. It has been shown that when the deoxygenation was applied in human red blood cells (RBCs), the anion exchange band 3 protein, which was recently identified as a potential substrate of PTP1B (51,52), was tyrosine-phosphorylated (53). Interestingly, under the deoxygenation, such as ischemic condition, the bioavailable level of NO is increased in RBCs (54), suggesting that PTP1B may be S-nitrosylated and therefore inactivated, concomitantly with an elevated tyrosine phosphorylation level of band 3.…”

Section: Discussionmentioning

confidence: 95%

Cysteine S-Nitrosylation Protects Protein-tyrosine Phosphatase 1B against Oxidation-induced Permanent Inactivation

Chen

Chu

Pan

et al. 2008

Journal of Biological Chemistry

139

120

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Protein S-nitrosylation mediated by cellular nitric oxide (NO) plays a primary role in executing biological functions in cGMPindependent NO signaling. Although S-nitrosylation appears similar to Cys oxidation induced by reactive oxygen species, the molecular mechanism and biological consequence remain unclear. We investigated the structural process of S-nitrosylation of protein-tyrosine phosphatase 1B (PTP1B). We treated PTP1B with various NO donors, including S-nitrosothiol reagents and compound-releasing NO radicals, to produce sitespecific Cys S-nitrosylation identified using advanced mass spectrometry (MS) techniques. Quantitative MS showed that the active site Cys-215 was the primary residue susceptible to S-nitrosylation. The crystal structure of NO donor-reacted PTP1B at 2.6 Å resolution revealed that the S-NO state at Cys-215 had no discernible irreversibly oxidized forms, whereas other Cys residues remained in their free thiol states. We further demonstrated that S-nitrosylation of the Cys-215 residue protected PTP1B from subsequent H 2 O 2 -induced irreversible oxidation. Increasing the level of cellular NO by pretreating cells with an NO donor or by activating ectopically expressed NO synthase inhibited reactive oxygen species-induced irreversible oxidation of endogenous PTP1B. These findings suggest that S-nitrosylation might prevent PTPs from permanent inactivation caused by oxidative stress.

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Section: Discussionmentioning

confidence: 95%

Cysteine S-Nitrosylation Protects Protein-tyrosine Phosphatase 1B against Oxidation-induced Permanent Inactivation

Chen

Chu

Pan

et al. 2008

Journal of Biological Chemistry

139

120

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“…Deoxygenation independently influenced both binding partners; specifically, differences between HbS and HbAo competition with GAPDH for cdB3 were apparent only when membranes isolated from deoxygenated RBCs were studied, suggesting that deoxygenation-dependent modification of cdB3 differentially influences affinity for (deoxy) HbS or Ao. On RBC deoxygenation, cdB3 is known to undergo phosphorylation (at multiple Tyr residues, possibly via p72 Syk activation or PTP1B inhibition) 34 and S-nitrosylation (at either/ both Cys 152 or Cys 330 ). 13 These modifications are well placed on cdB3 to inhibit either/both Hb and GAPDH binding, 22,29 as has been observed previously.…”

Section: Discussionmentioning

confidence: 99%

“…Competitive binding between GAPDH and Hb for cdB3: dependence on RBC O 2 content and cdB3 pTyr density cdB3 is known to undergo phosphorylation in a fashion that is linked to RBC O 2 content 34 and that diminishes GAPDH binding. 15 We confirmed this and found that O 2 content-responsive cdB3 phosphorylation in SSRBCs was significantly more robust than in normal RBCs (supplemental Figure 6A); however, increased cdB3pTyr in oxy-RBCs appeared insufficient to fully account for the substantial HMP constraint we observed in deoxy-RBCs by 1 H-NMR (supplemental Figure 6B).…”

Section: Quantitative Comparison Of Gapdh Binding To Bead-attached Mementioning

confidence: 99%

Sickle hemoglobin disturbs normal coupling among erythrocyte O2 content, glycolysis, and antioxidant capacity

Rogers¹,

Ross²,

Davignon

et al. 2013

Blood

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Key Points• Hb-conformation-dependent interaction with band 3 protein regulates glycolysis in RBCs.• In hypoxia, HbS disrupts this system, disabling RBC antioxidant defense. Energy metabolism in RBCs is characterized by O 2 -responsive variations in flux through the Embden Meyerhof pathway (EMP) or the hexose monophosphate pathway (HMP). Therefore, the generation of ATP, NADH, and 2,3-DPG (EMP) or NADPH (HMP) shift with RBC O 2 content because of competition between deoxyhemoglobin and key EMP enzymes for binding to the cytoplasmic domain of the Band 3 membrane protein (cdB3). Enzyme inactivation by cdB3 sequestration in oxygenated RBCs favors HMP flux and NADPH generation (maximizing glutathione-based antioxidant systems). We tested the hypothesis that sickle hemoglobin disrupts cdB3-based regulatory protein complex assembly, creating vulnerability to oxidative stress. In RBCs from patients with sickle cell anemia, we demonstrate in the present study constrained HMP flux, NADPH, and glutathione recycling and reduced resilience to oxidative stress manifested by membrane protein oxidation and membrane fragility. Using a novel, inverted membrane-on-bead model, we illustrate abnormal (O 2 -dependent) association of sickle hemoglobin to RBC membrane that interferes with sequestration/inactivation of the EMP enzyme GAPDH. This finding was confirmed by immunofluorescent imaging during RBC O IntroductionSickle cell anemia (SCA) arises from a single amino acid substitution (Glu6Val) in the ␤-globin chain. Although the change to hemoglobin (Hb) is simple and uniform, SCA is characterized by broad differences in clinical manifestation. Phenotype variation in SCA is thought to arise from both environmental and genetic factors (eg, ␤-gene cluster haplotype, degree of HbF expression, or effects of other epistatic genes). The environmental factor that most clearly influences SCA phenotype is hypoxia, which drives sickle Hb (HbS) polymerization and the resulting well-characterized alterations in RBC physiology and the microcirculation. However, the influence of hypoxia on the SCA phenotype appears to be insufficiently explained by HbS polymerization alone. 1 Moreover, we lack a clear mechanistic understanding of the significant oxidative stress complicating SCA, a key feature of phenotype variation, both at rest and in association with hypoxia. 2 Nonpolymerized, solution-phase HbS may promote oxidative stress, even in RBCs under normal physiologic O 2 gradients. 3 Specifically, the low redox potential for heme in HbS 4 and avid binding affinity of HbS for the cytoplasmic regulatory domain of the Band 3 membrane protein (cdB3) 5,6 strongly affect RBC energetics and antioxidant systems [7][8][9] and, notably, do so as a function of RBC O 2 content. Therefore, both the genesis and the disposal of reactive oxygen species are abnormal in SCA, creating a baseline state of oxidative stress, which worsens in hypoxia.In particular, consideration of metabolic control in RBCs suggests O 2 -dependent HbS-cdB3 interaction as a relatively ...

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“…3 An increase in band 3 tyrosine phosphorylation, which may reflect Syk activation or phosphotyrosine phosphatase (PTP) inhibition, was observed after vanadate or pervanadate treatment, 3,11-13 cell shrinkage induced by ionophore/Ca ϩϩ treatment or hypertonic conditions, 14,15 or elevation of intracellular Mg 2ϩ . 12 Deoxygenation also results in an increase in band 3 tyrosine phosphorylation in SS cells 2 and to a much lower extent in AA cells. 12 Evidence was provided recently that, on pervanadate treatment, band 3 is initially phosphorylated by Syk, providing docking sites (phosphotyrosines 8 and 21) for the binding of Lyn SH2 domain and subsequent phosphorylation of additional tyrosine residues (359 and 904).…”

mentioning

confidence: 97%

Deoxygenation of sickle cells stimulates Syk tyrosine kinase and inhibits a membrane tyrosine phosphatase

Merciris

Hardy‐Dessources

Giraud

2001

Blood

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Polymerization of hemoglobin S in sickle red cells, in deoxygenated conditions, is associated with K loss and cellular dehydration. It was previously reported that deoxygenation of sickle cells increases protein tyrosine kinase (PTK) activity and band 3 tyrosine phosphorylation and that PTK inhibitors reduce cell dehydration. Here, the study investigates which PTKs are involved and the mechanism of their activation. Deoxygenation of sickle cells induced a 2-fold increase in Syk activity, measured by autophosphorylation in immune complex assays, but had no effect on Lyn. Syk was not stimulated by deoxy-genation of normal red cells, and stimulation was partly reversible on reoxygen-ation of sickle cells. Syk activation was independent of the increase in intracellu-lar Ca and Mg 2 associated with deoxy-genation. Lectins that promote glyco-phorin or band 3 aggregation did not activate Syk. In parallel to Syk stimulation , deoxygenation of sickle cells, but not of normal red cells, decreased the activity of both membrane-associated protein tyrosine phosphatase (PTPs) and membrane protein thiol content. In vitro pretreatment of Syk immune complexes with membrane PTP inhibited Syk auto-phosphorylation. It is suggested that Syk activation in vivo could be mediated by PTP inhibition, itself resulting from thiol oxidation, as PTPs are known to be inhibited by oxidants. Altogether these data indicate that Syk could be involved in the mechanisms leading to sickle cell dehydration. (Blood. 2001;98:3121-3127)

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Deoxygenation and elevation of intracellular magnesium induce tyrosine phosphorylation of band 3 in human erythrocytes

Cited by 44 publications

References 21 publications

Cysteine S-Nitrosylation Protects Protein-tyrosine Phosphatase 1B against Oxidation-induced Permanent Inactivation

Cysteine S-Nitrosylation Protects Protein-tyrosine Phosphatase 1B against Oxidation-induced Permanent Inactivation

Sickle hemoglobin disturbs normal coupling among erythrocyte O2 content, glycolysis, and antioxidant capacity

Deoxygenation of sickle cells stimulates Syk tyrosine kinase and inhibits a membrane tyrosine phosphatase

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