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Fyn kinase regulates type II PtdIns 4-kinases in RBL 2H3 cells

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Abstract

Type II phosphatidylinositol 4-kinases are implicated in FcεRI-mediated signaling cascades leading to release of inflammatory molecules. Cross-linking of FcεRI on RBL 2H3 cells results in protein tyrosine phosphorylation and activation of type II PtdIns 4-kinase activity. Protein tyrosine kinase(s) that phosphorylate type II PtdIns 4-kinase(s) in RBL 2H3 cells remains elusive and is being addressed in this manuscript. Anti-Fyn kinase antibodies co-immunoprecipitated type II PtdIns 4-kinase activity from FcεRI cross-linked RBL 2H3 cells. In reciprocal assays, His-tagged types II PtdIns 4-kinases were shown to pull down Fyn kinase. Further, anti-Fyn immunoprecipitates were shown to phosphorylate type II PtdIns 4-kinase α and β in in vitro assays. Pull down studies with GST-Fyn-SH2 and GST-Fyn-SH3 domains showed that type II PtdIns 4-kinases associate with Fyn-SH2 domain. Knockdown of Fyn kinase in RBL 2H3 cells abrogated activation of type II PtdIns 4-kinase activity in response to FcεRI cross-linking and type II PtdIns 4-kinase activity in anti-phosphotyrosine immunoprecipitates. Knockdown of Fyn kinase was also strongly correlated with a reduction in β-hexosaminidase release in response to FcεRI cross-linking. These results suggest that type II PtdIns 4-kinases act downstream of Fyn kinase in FcεRI signaling cascades and are regulated by Fyn kinase.

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Acknowledgments

This research work was supported by grants in aid from Board of Research in Nuclear Sciences, Govt. of India (Sanction No. 2003/37/5/BRNS/667) and Council of Scientific and Industrial Research (sanction No. 37 (1099)02/EMR-II) to Dr. G. Subrahmanyam. Mr. Naveen B. and Ranjeet Kumar Sinha are supported by CSIR-SRF.

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  1. Naveen Bojjireddy

    Present address: Department of Anesthesiology and Pain Medicine, University of Washington, 850 Republican St, Room No: N130, Seattle, WA, 98109, USA

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  1. Department of Biosciences and Bioengineering, Wadhwani Research Centre for Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai, 400076, India

    Naveen Bojjireddy, Ranjeet Kumar Sinha & Gosukonda Subrahmanyam

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  1. Naveen Bojjireddy
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Correspondence to Naveen Bojjireddy.

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Bojjireddy, N., Sinha, R.K. & Subrahmanyam, G. Fyn kinase regulates type II PtdIns 4-kinases in RBL 2H3 cells. Mol Cell Biochem 387, 63–70 (2014). https://doi.org/10.1007/s11010-013-1871-8

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