Articles written in Journal of Biosciences
Volume 43 Issue 1 March 2018 pp 127-138 Article
Mast cells (MCs) respond to allergen challenge by release of pre-stored inflammatory mediators from their secretorygranules, on cross-linking of Fce receptor I (FceRI) receptors. The target-SNARE (t-SNARE) SNAP-23 has been shown toplay an important role in MC exocytosis and undergoes transient phosphorylation at Serine 95 (S95) and Serine 120 (S120),concomitant with mediator release. During current study we explored the importance of transient nature of phosphorylationat S120 in MC exocytosis. A phosphomimetic SNAP-23-S120D mutant of rodent SNAP-23 was cloned into EGFP vectorand its effect on the exocytosis and the mechanisms involved was studied in RBL-2H3 MC line. Secretion reporter assaywith SNAP-23-S120D transfected MCs revealed a very significant inhibition of exocytosis, and reduced ruffling inresponse to FceRI cross-linking. Further, the effect of this mutation on localization of SNAP-23 in MCs was studied.Immunofluorescence microscopy studies and membrane-cytosol fractionation of green fluorescent protein-tagged SNAP-23-S120D (GFP-SNAP-23-S120D) transfected MCs showed that a large proportion of GFP-SNAP-23-S120D was residingin cytosol unlike wild-type SNAP-23, in resting and activated MCs and even the membrane associated portion was oninternal lysosomal membranes than plasma membrane. These studies imply that dephosphorylation of S120 is important forSNAP-23 membrane association dynamics and subsequently MC degranulation.
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