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Fyn specifically Regulates the activity of red cell glucose-6-phosphate-dehydrogenase.

ABSTRACT: Fyn is a tyrosine kinase belonging to the Src family (Src-Family-Kinase, SFK), ubiquitously expressed. Previously, we report that Fyn is important in stress erythropoiesis. Here, we show that in red cells Fyn specifically stimulates G6PD activity, resulting in a 3-fold increase enzyme catalytic activity (kcat) by phosphorylating tyrosine (Tyr)-401. We found Tyr-401 on G6PD as functional target of Fyn in normal human red blood cells (RBC), being undetectable in G6PD deficient RBCs (G6PD-Mediterranean and G6PD-Genova). Indeed, Tyr-401 is located to a region of the G6PD molecule critical for the formation of the enzymatically active dimer. Amino acid replacements in this region are mostly associated with a chronic hemolysis phenotype. Using mutagenesis approach, we demonstrated that the phosphorylation status of Tyr401 modulates the interaction of G6PD with G6P and stabilizes G6PD in a catalytically more efficient conformation. RBCs from Fyn-/-mice are defective in G6PD activity, resulting in increased susceptibility to primaquine-induced intravascular hemolysis. This negatively affected the recycling of reduced Prx2 in response to oxidative stress, indicating that defective G6PD phosphorylation impairs defense against oxidation. In human RBCs, we confirm the involvement of the thioredoxin/Prx2 system in the increase vulnerability of G6PD deficient RBCs to oxidation. In conclusion, our data suggest that Fyn is an oxidative radical sensor, and that Fyn-mediated Tyr-401 phosphorylation, by increasing G6PD activity, plays an important role in the physiology of RBCs. Failure of G6PD activation by this mechanism may be a major limiting factor in the ability of G6PD deficient RBCs to withstand oxidative stress.

SUBMITTER: Alessandro M

PROVIDER: S-EPMC7387845 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Fyn specifically Regulates the activity of red cell glucose-6-phosphate-dehydrogenase.

Mattè Alessandro A Lupo Francesca F Tibaldi Elena E Di Paolo Maria Luisa ML Federti Enrica E Carpentieri Andrea A Pucci Piero P Brunati Anna Maria AM Cesaro Luca L Turrini Francesco F Gomez Manzo Saul S Choi Soo Young SY Marcial Quino Jaime J Kim Dae Won DW Pantaleo Antonella A Xiuli An A Iatcenko Iana I Cappellini Maria Domenica MD Forni Gian Luca GL De Franceschi Lucia L

Redox biology 20200711

Fyn is a tyrosine kinase belonging to the Src family (Src-Family-Kinase, SFK), ubiquitously expressed. Previously, we report that Fyn is important in stress erythropoiesis. Here, we show that in red cells Fyn specifically stimulates G6PD activity, resulting in a 3-fold increase enzyme catalytic activity (k<sub>cat</sub>) by phosphorylating tyrosine (Tyr)-401. We found Tyr-401 on G6PD as functional target of Fyn in normal human red blood cells (RBC), being undetectable in G6PD deficient RBCs (G6P ...[more]

PMID: 32863204

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Project description:Neonatal jaundice is a common and severe disease in premature infants with Glucose-6-Phosphate Dehydrogenase (G-6-PD) deficiency. The World Health Organization (WHO) has recommended screening for G-6-PD deficiency in newborns for early recognition as well as to prevent unwanted outcomes in a timely manner. The present study aimed to assess a point-of-care, careSTARTTM G6PD biosensor as a quantitative method for the diagnosis of G-6-PD deficiency. Factors influencing the evaluation of G-6-PD enzyme activity were examined in 40 adults, including ethylenediaminetetraacetic acid (EDTA) anticoagulant, hematocrit concentration, storage temperature and time. Analytic performance of the careSTARTTM G6PD biosensor was evaluated in 216 newborns and compared with fluorescent spot test (FST) and standard quantitative G-6-PD enzyme activity (SGT) assay. The results of factors affecting the G-6-PD enzyme activity showed that the activity determined from finger-prick was not statistically different from venous blood (p = 0.152). The G-6-PD value was highly dependent on the hematocrit and rose with increasing hematocrit concentration. Its activity was stable at 4°C for 3 days. Reliability analysis between the careSTARTTM G6PD biosensor and SGT assay showed a strong correlation with a Pearson's correlation coefficient of 0.82 and perfect agreement by intraclass correlation coefficient (ICC) of 0.90. Analysis of the area under the Receiver Operating Curve (AUC) illustrated that the careSTARTTM G6PD biosensor had 100% sensitivity, 96% specificity, 73% positive predictive value (PPV), 100% negative predictive value (NPV) and 97% accuracy at 30% of residual activity. While the diagnostic ability for identifying G-6-PD deficiency had 78% sensitivity, 89% specificity, 56% positive predictive value (PPV), 96% negative predictive value (NPV) and 88% accuracy when stratified by gender. The careSTARTTM G6PD biosensor is an attractive option as a point-of-care quantitative method for G-6-PD activity detection. Quantification of G-6-PD enzyme activity in newborns is the most effective approach for the management of G-6-PD deficiency to prevent severe jaundice and acute hemolysis.

Dataset Information

Fyn specifically Regulates the activity of red cell glucose-6-phosphate-dehydrogenase.

Publications

Fyn specifically Regulates the activity of red cell glucose-6-phosphate-dehydrogenase.

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