Regulation of MT1-MMP Activity through Its Association with ERMs.

M Victoria Hernández-Riquer,Soraya López-Martín,Henar Suárez,Víctor Toribio,Laura Genís,María Yáñez-Mó,Alicia G Arroyo,Moreno Zamai

doi:10.3390/cells9020348

Abstract

Membrane-bound proteases play a key role in biology by degrading matrix proteins or shedding adhesion receptors. MT1-MMP metalloproteinase is critical during cancer invasion, angiogenesis, and development. MT1-MMP activity is strictly regulated by internalization, recycling, autoprocessing but also through its incorporation into tetraspanin-enriched microdomains (TEMs), into invadopodia, or by its secretion on extracellular vesicles (EVs). We identified a juxtamembrane positively charged cluster responsible for the interaction of MT1-MMP with ERM (ezrin/radixin/moesin) cytoskeletal connectors in breast carcinoma cells. Linkage to ERMs regulates MT1-MMP subcellular distribution and internalization, but not its incorporation into extracellular vesicles. MT1-MMP association to ERMs and insertion into TEMs are independent phenomena, so that mutation of the ERM-binding motif in the cytoplasmic region of MT1-MMP does not preclude its association with the tetraspanin CD151, but impairs the accumulation and coalescence of CD151/MT1-MMP complexes at actin-rich structures. Conversely, gene deletion of CD151 does not impact on MT1-MMP colocalization with ERM molecules. At the plasma membrane MT1-MMP autoprocessing is severely dependent on ERM association and seems to be the dominant regulator of the enzyme collagenolytic activity. This newly characterized MT1-MMP/ERM association can thus be of relevance for tumor cell invasion.

Highlights

Extracellular matrix (ECM) hydrolysis and rupture is performed by a large group of matrix metalloproteinases (MMPs), which can be either secreted into the extracellular milieu or transmembrane proteins
To assess whether this is the case for MT1-MMP, we performed an enzyme-linked immunosorbent assay (ELISA) in vitro binding assay using synthetic peptides encoding the C-terminal sequence of MT1-MMP and the recombinant N-terminal domain of moesin fused to GST
Our results demonstrated the interaction between wildtype MT1-MMP and moesin in vitro, that was completely abrogated by mutation of the juxtamembrane RRH563 cluster (Figure 1A)

Summary

Introduction

Extracellular matrix (ECM) hydrolysis and rupture is performed by a large group of matrix metalloproteinases (MMPs), which can be either secreted into the extracellular milieu or transmembrane proteins. Since evidence showed a strong correlation between MT1-MMP expression levels and tumour invasiveness [12,13,14,15,16], a lot of research has been focused in unravelling the mechanisms that control its activity. Due to the variability of substrates cleaved by this enzyme and the low expression levels needed for detecting a significant effect, MT1-MMP activity has to be tightly regulated. Processing of two main substrates of MT1-MMP, collagen and pro-MMP2, requires dimerization of the protease [18] through its hemopexin domain. This dimerization probably immobilizes the protein, facilitating its action over the collagen triple helix.

Methods

Results

Conclusion