Kallikrein-Related Peptidase 14 Activates Zymogens of Membrane Type Matrix Metalloproteinases (MT-MMPs)—A CleavEx Based Analysis

Abstract

Kallikrein-related peptidases (KLKs) and matrix metalloproteinases (MMPs) are secretory proteinases known to proteolytically process components of the extracellular matrix, modulating the pericellular environment in physiology and in pathologies. The interconnection between these families remains elusive. To assess the cross-activation of these families, we developed a peptide, fusion protein-based exposition system (Cleavage of exposed amino acid sequences, CleavEx) aiming at investigating the potential of KLK14 to recognize and hydrolyze proMMP sequences. Initial assessment identified ten MMP activation domain sequences which were validated by Edman degradation. The analysis revealed that membrane-type MMPs (MT-MMPs) are targeted by KLK14 for activation. Correspondingly, proMMP14-17 were investigated in vitro and found to be effectively processed by KLK14. Again, the expected neo-N-termini of the activated MT-MMPs was confirmed by Edman degradation. The effectiveness of proMMP activation was analyzed by gelatin zymography, confirming the release of fully active, mature MT-MMPs upon KLK14 treatment. Lastly, MMP14 was shown to be processed on the cell surface by KLK14 using murine fibroblasts overexpressing human MMP14. Herein, we propose KLK14-mediated selective activation of cell-membrane located MT-MMPs as an additional layer of their regulation. As both, KLKs and MT-MMPs, are implicated in cancer, their cross-activation may constitute an important factor in tumor progression and metastasis.

Matrix Metalloproteinases Shape the Tumor Microenvironment in Cancer Progression

“…The other membrane-bound MMPs -15, -17, -24, and -25 also contain a furin cleavage site and are believed to be activated in a similar manner prior to their membrane association [ 84 ]. Kallikrein-related peptidases can also proteolytically activate proMMPs [ 85 ].…”

DOT1L stimulates MYC/Mondo transcription factor activity by promoting its degradation cycle on chromatin

“…To facilitate cleavage detection, we developed a reporter construct using the Porphyromonas gingivalis HmuY protein as a scaffold due to its protease-resistant nature. 96,97 Since this protein has been used in CleavEx 98 in vitro technology, but not yet in cells, we first tested its expression in the HCT116 and U2OS cell lines. The protease-resistant HmuY protein was expressed strongly in HCT116 cells (Supplemental Figure 6D) .…”

Comparative analyses of Netherton syndrome patients and Spink5 conditional knock-out mice uncover disease-relevant pathways

“…Interestingly, loss of CD34+ fibroblasts has been observed in other inflammatory skin diseases 42 . The down-regulation of collagens and CD34 could also be related to increased matrix-metalloproteinase and KLK activities in lesional skin 43 and implies a possible hair growth defect (caused by depletion of CD34+ hair follicle progenitor cells) in addition to hair shaft abnormalities described in NS 44 .…”