Modulation of RECK levels in Xenopus A6 cells: effects on MT1-MMP, MMP-2 and pERK levels

Jessica A Willson,Sashko Damjanovski,Bradley S Bork,Carlie A Muir

doi:10.1186/s40709-019-0108-8

Jessica A Willson, Sashko Damjanovski + Show 2 more

Open Access

https://doi.org/10.1186/s40709-019-0108-8

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Abstract

BackgroundMT1-MMP is a cell-surface enzyme whose regulation of pro-MMP-2 and ERK activation position it as a key facilitator of ECM remodelling and cell migration. These processes are modulated by endogenous MMP inhibitors, such as RECK, a GPI-anchored protein which has been shown to inhibit both MT1-MMP and MMP-2 activity. Our previous studies have revealed a link between MT1-MMP levels, and pro-MMP-2 and ERK activation in mammalian cells, as well as MT1-MMP and RECK co-localization in Xenopus embryos. We here investigated how modulation of RECK would impact MT1-MMP and MMP-2 levels, as well as ERK signalling in Xenopus A6 cells.ResultsWe used a Morpholino approach to knockdown RECK, plasmid transfection to overexpress RECK, and PI-PLC treatment to shed RECK from the cell surface of Xenopus A6 cells. RECK reduction did not alter pERK or MT1-MMP levels, nor MMP-2 activity as measured by zymography; thus RECK-knockdown cells maintained the ability to remodel the ECM. RECK overexpression and PI-PLC treatment both increased ECM remodelling potential through increased MT1-MMP protein and relative MMP-2 activation levels.ConclusionsRECK changes that reduce the ability of the cell to remodel the ECM (overexpression and cell surface shedding) are compensated for by increases in MT1-MMP, and MMP-2 levels as seen by zymography.

Highlights

MT1-matrix metalloproteinases (MMPs) is a cell-surface enzyme whose regulation of pro-MMP-2 and ERK activation position it as a key facilitator of extracellular matrix (ECM) remodelling and cell migration
Having confirmed the efficacy of the RECK MO, we examined the levels of MMPs and their activity in A6 cells following RECK knockdown
To address if the maintenance of low levels of MMPs is a common mechanism seen in non-mammalian epithelial cells, here we investigate the impact of altered RECK levels in the context of the ECM remodelers MT1MMP and MMP-2, as well as ERK in Xenopus A6 cells

Summary

Introduction

MT1-MMP is a cell-surface enzyme whose regulation of pro-MMP-2 and ERK activation position it as a key facilitator of ECM remodelling and cell migration These processes are modulated by endogenous MMP inhibitors, such as RECK, a GPI-anchored protein which has been shown to inhibit both MT1-MMP and MMP-2 activity. In mammalian MDA-MB-231 cells, inhibition of pro-MT1-MMP activation elevated global MT1MMP and pERK levels, but decreased MMP-2 levels [12] These mammalian cell studies demonstrated that when MT1-MMP levels are altered, pERK and MMP-2 activation levels alter in different ways depending on treatment. This suggests a mechanism that may involve more molecules than just MT1-MMP in the activation of ERK and pro-MMP-2, with RECK possibly playing an important role. Several other studies have described interactions between RECK and MT1-MMP proteins in vivo and in vitro [15,16,17], with RECK being shown to complex with MT1-MMP at the cell surface to both attenuate its proteolytic activity and modulate its endocytosis from the cell surface [18]

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