Spatial analysis of RECK, MT1-MMP, and TIMP-2 proteins during early Xenopus laevis development

Willson, Jessica A.; Damjanovski, Sashko

doi:10.1016/j.gep.2019.119066

Cited by 3 publications

(1 citation statement)

References 34 publications

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“…We have previously cloned and characterized the RECK gene and studied its expression during Xenopus laevis development, showing that RECK is present at stages where ECM remodeling events are associated with neural function [13]. Our recent in vivo examination of RECK, MT1-MMP, and TIMP-2 show that these proteins colocalize in the dorsal axis of 48-h Xenopus tailbud stage embryos, particularly in the neural tube [14]. Several other studies have also described interactions between RECK and MT1-MMP proteins in vivo and in vitro [15–17], 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].…”

Section: Introductionmentioning

confidence: 99%

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

Willson

Bork

Muir

et al. 2019

J of Biol Res-Thessaloniki

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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.

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Section: Introductionmentioning

confidence: 99%

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

Willson

Bork

Muir

et al. 2019

J of Biol Res-Thessaloniki

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Post-translational activation of Mmp2 correlates with patterns of active collagen degradation during the development of the zebrafish tail

Wyatt

Crawford

2021

Developmental Biology

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Analysis of the inhibiting activity of reversion-inducing cysteine-rich protein with Kazal motifs (RECK) on matrix metalloproteinases

Mendes

Amo-Maestro

Marino-Puertas

et al. 2020

Sci Rep

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Matrix metalloproteinases (MMPs) occur in 23 human paralogues with key functions in physiology, and their activity is controlled by protein inhibitors. Reversion-inducing cysteine-rich protein with Kazal motifs (RecK), which is essential for embryogenesis and tumour suppression, has been reported to inhibit MMPs. Here, we developed eukaryotic and bacterial expression systems for different RECK variants and analysed their inhibitory capacity against representative MMps in vitro. We could not detect any significant inhibition. Instead, we found that partially purified RECK from the conditioned medium of transfected Expi293F cells but not that of ExpiCHO-S or Drosophila Schneider cells contained a contaminant with proteolytic activity. the contaminant was removed through treatment with a small-molecule serine peptidase inhibitor and additional chromatographic purification. A tantamount contaminant was further detected in an equivalent expression system of the n-terminal fragment of the proteoglycan testican 3, but not in those of two other proteins. These results indicate that previous reports of inhibitory activity of recombinant RecK on MMps, which were performed with partially purified samples, were probably masked by a coeluting contaminant present in the supernatant of HEK293-derived cells. Thus, RECK is probably not a direct inhibitor of MMP catalytic activity but may still regulate MMps through other mechanisms. Proteolysis is a post-translational modification of proteins and peptides that is essential for all physiological pathways. It is exerted by peptidases, among which metallopeptidases (MPs) are one of several chemical classes and consist of various clans and families 1. The matrix metalloproteinases (MMPs; 23 paralogues in humans), as well as the ADAMs/adamalysins (19 in humans) and the more distantly related ADAMTSs (19 in humans) are among the most studied MPs because of their enormous relevance for human health and disease 2-8. Collectively, they carry out functions as broad degraders during the digestion of intake proteins, turnover of extracellular-matrix components for tissue remodelling and developmental processes, and clearance of obsolete or malfunctioning polypeptides. Moreover, they are fine regulators of shedding, maturation and inactivation of other proteins through limited proteolysis of one or a few peptide bonds 9. Peptide-bond scission is normally irreparable under physiological conditions, so MPs must be fastidiously controlled to avoid aberrant cleavage that would cause dysfunction and pathology. This regulation is physiologically carried out for MMPs in humans by four tissue inhibitors of metalloproteinases and the broad-spectrum pan-peptidase inhibitor α 2-macroglobulin 3,10-12. Another reported inhibitor is the protein RECK 13-17. RECK, an acronym for reversion-inducing cysteine-rich protein with Kazal motifs, is encoded by a gene that suppresses the transformed phenotype caused by ras oncogenes 13,18. The 971-residue molecule is a membrane-anchored glycoprotein of ~125 kDa, which...

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Spatial analysis of RECK, MT1-MMP, and TIMP-2 proteins during early Xenopus laevis development

Cited by 3 publications

References 34 publications

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

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

Post-translational activation of Mmp2 correlates with patterns of active collagen degradation during the development of the zebrafish tail

Analysis of the inhibiting activity of reversion-inducing cysteine-rich protein with Kazal motifs (RECK) on matrix metalloproteinases

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