Regulator of G-Protein Signalling-14 (RGS14) Regulates the Activation of αMβ2 Integrin during Phagocytosis.

Jenson Lim,Neil A Hotchin,Robin C May,Jo Thompson,Emmanuelle Caron,Jon C.D Houtman

doi:10.1371/journal.pone.0069163

Jenson Lim, Neil A Hotchin + Show 4 more

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https://doi.org/10.1371/journal.pone.0069163

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Abstract

Integrin-mediated phagocytosis, an important physiological activity undertaken by professional phagocytes, requires bidirectional signalling to/from αMβ2 integrin and involves Rap1 and Rho GTPases. The action of Rap1 and the cytoskeletal protein talin in activating αMβ2 integrins, in a RIAM-independent manner, has been previously shown to be critical during phagocytosis in mammalian phagocytes. However, the events downstream of Rap1 are not clearly understood. Our data demonstrate that one potential Rap1 effector, Regulator of G-Protein Signalling-14 (RGS14), is involved in activating αMβ2. Exogenous expression of RGS14 in COS-7 cells expressing αMβ2 results in increased binding of C3bi-opsonised sheep red blood cells. Consistent with this, knock-down of RGS14 in J774.A1 macrophages results in decreased association with C3bi-opsonised sheep red blood cells. Regulation of αMβ2 function occurs through the R333 residue of the RGS14 Ras/Rap binding domain (RBD) and the F754 residue of β2, residues previously shown to be involved in binding of H-Ras and talin1 head binding prior to αMβ2 activation, respectively. Surprisingly, overexpression of talin2 or RAPL had no effect on αMβ2 regulation. Our results establish for the first time a role for RGS14 in the mechanism of Rap1/talin1 activation of αMβ2 during phagocytosis.

Highlights

Phagocytosis has a critical physiological function as part of the feeding process in amoebae, or as part of the innate immune system which functions to remove microorganisms and apoptotic cells in mammals [1]
In humans the process of phagocytosis is undertaken by professional phagocytes such as neutrophils and macrophages
Sheep red blood cells (RBC) were purchased from TCS Biosciences, Ltd., Gelatin Veronal Buffer, and C5-deficient serum were from Sigma

Summary

Introduction

Phagocytosis has a critical physiological function as part of the feeding process in amoebae, or as part of the innate immune system which functions to remove microorganisms and apoptotic cells in mammals [1]. Phagocytosis involves receptor-mediated particle recognition, actin-driven uptake, phagosome maturation and particle clearance. Surfaceexpressed phagocytic receptors exist that can bind their target directly or indirectly through opsonins, depending on cell type and the nature of its targets [2]. Two well characterized phagocytic receptors are the Fcγ receptor (FcγR) and complement receptor 3 (aka CR3, Mac-1, αMβ2, CD11b/ CD18), that bind IgG- or C3bi-opsonised particles, respectively [1,2]. Integrin subunits are made up of a large extracellular ligand-binding domain, a single pass transmembrane domain and a short cytoplasmic tail and they are bi-directionally regulated. Association with a complement-opsonised particle leads to “outside-in” signalling which leads to the RhoAdependent uptake, and subsequent destruction, of the particle [7,8]

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