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KMID : 0381120190410020241
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Genes and Genomics
2019 Volume.41 No. 2 p.241 ~ p.248
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Roles of paxillin phosphorylation in IL-3 withdrawal-induced Ba/F3 cell apoptosis
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Nah Ae-Sun
Chay Kee-Oh
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Abstract
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Background: Ba/F3, a mouse pro-B cell line, is dependent on IL-3 for its survival and proliferation. IL-3 withdrawal causes cells to round, stop in G1 phase, then undergo apoptosis. Additionally, IL-3 is known to induce tyrosine phosphorylation of paxillin, a scaffold and signaling protein. We previously determined that overexpression of paxillin prohibited Ba/F3 cell apoptosis induced by IL-3 withdrawal.
Objective: Address whether phosphorylation is essential for the anti-apoptotic effect of overexpressed paxillin.
Methods: Mutations were introduced into paxillin cDNA at five phosphorylation sites?Y31F, Y40F, Y118F, Y181F, S273A, or S273D. After overexpression of paxillin mutants in Ba/F3 cells, the apoptotic proportions of cell populations were measured by an annexin V conjugation assay while cells were undergoing IL-3 withdrawal.
Results: The anti-apoptotic effect of paxillin overexpression was abolished by site-directed mutagenesis replacing Y31, Y40, Y118, and Y181 with phenylalanine, and S273 with aspartic acid. In contrast, the mutation replacing S273 with alanine had no effect on the anti-apoptotic effect.
Conclusion: The above results suggest that paxillin-mediated phosphorylation at Y31, Y40, Y118, and Y181 is essential for the anti-apoptotic effect of paxillin overexpression in Ba/F3 cells and contributes to the cell survival signaling pathway triggered by IL-3. Conversely, phosphorylation at S273 is involved in the negative regulation of the anti-apoptotic action of overexpressed paxillin.
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KEYWORD
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Paxillin, Phosphorylation, Survival, Apoptosis, IL-3, B cell
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