Topographical interrogation of the living cell surface reveals its role in rapid cell shape changes during phagocytosis and spreading

Maha A Al Jumaa,Sharon Dewitt,Maurice B Hallett

doi:10.1038/s41598-017-09761-6

Maha A Al Jumaa, Sharon Dewitt + Show 1 more

Open Access

https://doi.org/10.1038/s41598-017-09761-6

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Journal: Scientific Reports	Publication Date: Aug 29, 2017
Citations: 15	License type: open-access

Affiliation: Cardiff University

Abstract

Dramatic and rapid changes in cell shape are perhaps best exemplified by phagocytes, such as neutrophils. These cells complete the processes of spreading onto surfaces, and phagocytosis within 100 s of stimulation. Although these cell shape changes are accompanied by an apparent large increase in cell surface area, the nature of the membrane “reservoir” for the additional area is unclear. One proposal is that the wrinkled cell surface topography (which forms micro-ridges on the neutrophil surface) provides the resource for neutrophils to expand their available surface area. However, it has been problematic to test this proposal in living cells because these surface structures are sub-light microscopic. In this paper, we report the development of a novel approach, a variant of FRAP (fluorescent recovery after photo-bleaching) modified to interrogate the diffusion path-lengths of membrane associated molecules. This approach provides clear evidence that the cell surface topography changes dramatically during neutrophil shape change (both locally and globally) and can be triggered by elevating cytosolic Ca2+.

Highlights

The lipid bilayers that make up the plasma membrane of living cells are strong transversely, but are intrinsically weak in the lateral direction and cannot stretch significantly without rupture[1]
In this paper we have developed a novel approach, which allows information of the cell surface topography to be extracted from living cells, and so permits dynamic changes in cell surface topography to be followed
The effect of the surface terrain on the apparent D constant has been re-evaluated[25] and modelling suggests that the relationship between the apparent D constant and the surface over which it is measured is relevant

Summary

Introduction

The lipid bilayers that make up the plasma membrane of living cells are strong transversely (ie across the water-lipid-water sandwich), but are intrinsically weak in the lateral direction and cannot stretch significantly without rupture[1]. A biophysical approach has shown that the wrinkled surface can be unwrinkled by applying suction through a micropipette, and that the force required to do so is reduced during phagocytosis[8, 9] This points to a slackening of the forces holding the wrinkles in place during triggering of neutrophil shape change. Topographical deviation from the planar would increase the actual 2D path length (Fig. 1) This difference in timing reflects the smoothness or wrinkledness of the path-length for diffusion. Using this approach, we show that non-spread living neutrophils have significant surface wrinkledness, which is lost (i) at the spread uropod tail during chemotaxis and (ii) locally near the phagocytic cup during phagocytosis, around the phagosome and extending pseudopodia. The surface topography can be altered experimentally by osmotically active media; by membrane expanders and by IP3-triggered Ca2+ influx, the physiologically relevant trigger[10,11,12,13]

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