Calcium-dependent, non-apoptotic, large plasma membrane bleb formation in physiologically stimulated mast cells and basophils

C Jansen,C Tobita,E U Umemoto,J Starkus,N M Rysavy,L M N Shimoda,C Sung,A.J Stokes,H Turner

doi:10.1080/20013078.2019.1578589

C Jansen, C Tobita + Show 7 more

Open Access

https://doi.org/10.1080/20013078.2019.1578589

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Abstract

ABSTRACTLarge membrane derangements in the form of non-detaching blebs or membrane protrusions occur in a variety of cell stress and physiological situations and do not always reflect apoptotic processes. They have been studied in model mast cells under conditions of cell stress, but their potential physiological relevance to mast cell function and formation in primary mast cells or basophils have not been addressed. In the current study, we examine the large, non-detaching, non-apoptotic, membrane structures that form in model and primary mast cells under conditions of stimulation that are relevant to allergy, atopy and Type IV delayed hypersensitivity reactions. We characterized the inflation kinetics, dependency of formation upon external free calcium and striking geometric consistency of formation for large plasma membrane blebs (LPMBs). We describe that immunologically stimulated LPMBs in mast cells are constrained to form in locations where dissociation of the membrane-associated cytoskeleton occurs. Mast cell LPMBs decorate with wheat germ agglutinin, suggesting that they contain plasma membrane (PM) lectins. Electrophysiological capacitance measurements support a model where LPMBs are not being formed from internal membranes newly fused into the PM, but rather arise from stretching of the existing membrane, or inflation and smoothing of a micro-ruffled PM. This study provides new insights into the physiological manifestations of LPMB in response to immunologically relevant stimuli and in the absence of cell stress, death or apoptotic pathways.

Highlights

Large-diameter, low-curvature membrane derangements have been described in a variety of cell stress and cellular activation situations
Cells were stimulated with IgE alone and with IgE anti-DNP followed by antigenic crosslinking with keyhole limpet hemocyanin (KLH)-DNP to induce FcεRI signal transduction
large plasma membrane blebs (LPMBs) are enigmatic structures, and the present study provides new data on their manifestation in mast cells

Summary

Introduction

Large-diameter, low-curvature membrane derangements have been described in a variety of cell stress and cellular activation situations. Large-diameter, lowcurvature membrane protrusions that can be induced by treatments such as pulsed laser exposure and chemicals such as dithiothreitol and paraformaldehyde [1] have previously been termed giant plasma membrane vesicles (GPMVs) and been used as tools to assess fluid/fluid phase partitioning in membranes and the association of receptors with particular lipid micro-domains [2,3,4]. Large membrane protrusions that resemble GPMV have been associated with cellular activation, mitosis, locomotion and apoptosis [5,6,7], and are more accurately termed as blebs due to their non-detaching and non-vesicular nature. A rat basophilic leukaemia cell line that is a common mast cell model has been used (primarily as a system of convenience) to generate these protrusions for lipid micro-domain studies. The non-apoptotic presentation of such protrusions has been understudied and may represent a poorly understood process in activation of mast cells in response to physiological stimuli

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