Non-proteolytic, receptor/ligand interactions associate cellular membrane type-1 matrix metalloproteinase with the complement component C1q

Membrane type-1 matrix metalloproteinase (MT1-MMP), a prototypic member of the membrane-tethered MMP family, is an essential component of a cellular proteolysis apparatus. Recognition of protein cleavage targets followed by proteolysis is a main function of MT1-MMP. For the first time, however, we present evidence that MT1-MMP and other structurally related membrane MMPs bind C1q, the recognition unit of the first component of complement C1 that initiates activation of the classical pathway of complement. These interactions involve the catalytic domain of MT1-MMP and the C1q globular domain. In silico modeling followed by mutagenesis and the in vitro and cell-based binding studies showed that the His(171)-Glu-Lys-Gln-Ala-Asp(176) and Val(223)-Arg-Asn(224) peptide sequences of MT1-MMP are directly involved in the binding with C1q. These sequence regions are spatially distant from the active site of the protease. As a result, the catalytically active and the catalytically latent forms of cellular MT1-MMP are both efficient in binding with C1q. In agreement, despite the MT1-MMP/C1q interactions, C1q is totally resistant to MT1-MMP proteolysis. The discovery of the unconventional, receptor/ligand-like interactions of MT1-MMP with C1q, an essential component of immunity, is a significant step toward a more complete understanding of the role of this membrane-tethered protease in cancer.     

The hemopexin-like C-terminal domain of membrane type 1 matrix metalloproteinase regulates proteolysis of a multifunctional protein, gC1qR.

Matrix metalloproteinases (MMPs) including membrane type 1 MMP (MT1-MMP) can degrade extracellular matrix and cell surface receptor molecules and have an essential function in malignancy. Recently, we established a functional link between MT1-MMP and the receptor of complement component 1q (gC1qR). The gC1qR is known as a compartment-specific regulator of diverse cellular and viral proteins. Once released by proliferating cells, soluble gC1qR may inhibit complement component 1q hemolytic activity and play important roles in vivo in assisting tumor cells to evade destruction by complement. Here, we report that gC1qR is susceptible to MT1-MMP proteolysis in vitro and in cell cultures. The major MT1-MMP cleavage site (Gly(79) down arrow Gln(80)) is localized within the structurally disordered loop connecting the beta(3) and the beta(4) strands of gC1qR. The recombinant MT1-MMP construct that included the catalytic domain but lacked the hemopexin-like domain lost the proteolytic capacity; however, it retained the ability to bind gC1qR. Inhibition of MT1-MMP activity by a hydroxamate inhibitor converted the protease into a cell surface receptor of gC1qR and promoted co-precipitation MT1-MMP with the soluble gC1qR protein. It is tempting to hypothesize that these novel mechanisms may play important roles in vivo and have to be taken into account in designing hydroxamate-based cancer therapy.     

The low density lipoprotein receptor-related protein LRP is regulated by membrane type-1 matrix metalloproteinase (MT1-MMP) proteolysis in malignant cells

We demonstrate that the presentation of LRP and the subsequent uptake of its ligands by malignant cells are both strongly regulated by MT1-MMP. Because LRP is essential for the clearance of multiple ligands, these findings have important implications for many pathophysiological processes including the pericellular proteolysis in neoplastic cells as well as the fate of the soluble matrix-degrading proteases such as MMP-2. MT1-MMP is a key protease in cell invasion and a physiological activator of MMP-2. Cellular LRP consists of a non-covalently associated 515-kDa extracellular alpha-chain (LRP-515) and an 85-kDa membrane-spanning beta-chain, and plays a dual role as a multifunctional endocytic receptor and a signaling molecule. Through the capture and uptake of several soluble proteases, LRP is involved in the regulation of matrix proteolysis. LRP-515 associates with the MT1-MMP catalytic domain and is highly susceptible to MT1-MMP proteolysis in vitro. Similar to MT1-MMP, the metalloproteinases MT2-MMP, MT3-MMP and MT4-MMP also degrade LRP. The N-terminal and C-terminal parts of the LRP-515 subunit are resistant and susceptible, respectively, to MT1-MMP proteolysis. In cells co-expressing LRP and MT1-MMP, the proteolytically competent protease decreases the levels of cellular LRP and releases its N-terminal portion in the extracellular milieu while the catalytically inert protease co-precipitates with LRP. These events implicate MT1-MMP, not only in the activation of MMP-2, but also in the mechanisms that control the subsequent fate of MMP-2 in cells and tissues.     

Matrix-dependent proteolysis of surface transglutaminase by membrane-type metalloproteinase regulates cancer cell adhesion and locomotion

Cell invasion requires cooperation between adhesion receptors and matrix metalloproteinases (MMPs). Membrane type (MT)-MMPs have been thought to be primarily involved in the breakdown of the extracellular matrix. Our report presents evidence that MT-MMPs in addition to the breakdown of the extracellular matrix may be engaged in proteolysis of adhesion receptors on tumor cell surfaces. Overexpression of MT1-MMP by glioma and fibrosarcoma cells led to proteolytic degradation of cell surface tissue transglutaminase (tTG) at the leading edge of motile cancer cells. In agreement, structurally related MT1-MMP, MT2-MMP, and MT3-MMP but not evolutionary distant MT4-MMP efficiently degraded purified tTG in vitro. Because cell surface tTG represents a ubiquitously expressed, potent integrin-binding adhesion coreceptor involved in the binding of cells to fibronectin (Fn), the proteolytic degradation of tTG by MT1-MMP specifically suppressed cell adhesion and migration on Fn. Reciprocally, Fn in vitro and in cultured cells protected its surface receptor, tTG, from proteolysis by MT1-MMP, thereby supporting cell adhesion and locomotion. In contrast, the proteolytic degradation of tTG stimulated migration of cells on collagen matrices. Together, our observations suggest both an important coreceptor role for cell surface tTG and a novel regulatory function of membrane-anchored MMPs in cancer cell adhesion and locomotion. Proteolysis of adhesion proteins colocalized with MT-MMPs at discrete regions on the surface of migrating tumor cells might be controlled by composition of the surrounding ECM.     

Cell-surface-associated tissue transglutaminase is a target of MMP-2 proteolysis

MT1-MMP, a prototypic member of a membrane-type metalloproteinase subfamily, is an invasion promoting protease and an activator of MMP-2. In addition, MT1-MMP proteolysis regulates the functionality of cell-surface adhesion/signaling receptors including tissue transglutaminase (tTG). tTG is known to serve as an adhesion coreceptor for beta1/beta3 integrins and as an enzyme that catalyzes the cross-linking of proteins and the conjugation of polyamines to proteins. Here, we report that MMP-2, functioning in concert with MT1-MMP, hydrolyzes cell-surface-associated tTG, thereby further promoting the effect initiated by the activator of MMP-2. tTG, in return, preferentially associates with the activation intermediate of MMP-2. This event decreases the rate of MMP-2 maturation and protects tTG against proteolysis by MMP-2. Our cell culture, in vitro experiments, and in silico modeling indicate that the catalytic domain of MMP-2 directly associates with the core enzymatic domain II of tTG (the K(d) = 380 nM). The follow-up cleavage of the domain II eliminates both the receptor and the enzymatic activity of tTG. Our data illuminate the coordinated interplay involving the MT1-MMP/MMP-2 protease tandem in the regulation of the cell receptors and explain the underlying biochemical mechanisms of the extensive tTG proteolysis that exists at the normal tissue/tumor boundary. Our findings also suggest that neoplasms, which express functionally active MT1-MMP and, therefore, activate soluble MMP-2, can contribute to the degradation of tTG expressed in neighboring host cells. The loss of adhesive and enzymatic activities of tTG at the interface between tumor and normal tissue will decrease cell-matrix interactions and inhibit matrix cross-linking, causing multiple pathological alterations in host cell adhesion and locomotion.     

The type I collagen induction of MT1-MMP-mediated MMP-2 activation is repressed by alphaVbeta3 integrin in human breast cancer cells.

The influence of alphaVbeta3 integrin on MT1-MMP functionality was studied in human breast cancer cells of differing beta3 integrin status. Overexpression of beta3 integrin caused increased cell surface expression of alphaV integrin and increased cellular adhesion to extracellular matrix (ECM) substrates in BT-549, MDA-MB-231 and MCF-7 cells. beta3 integrin expression also enhanced the migration of breast cancer cells on ECM substrates and enhanced collagen gel contraction. In vivo, alphaVbeta3 cooperated with MT1-MMP to increase the growth of MCF-7 cells after orthotopic inoculation in immunocompromised mice, but had no influence on in vitro proliferation. Despite these stimulatory effects, overexpression of beta3 integrin suppressed the type I collagen (Col I) induced MMP-2 activation in all breast cancer cell lines analyzed. This was also evident in extracts from the MCF-7 tumors in vivo, where MMP-2 activation was stimulated by MT1-MMP transfection, but attenuated with beta3 integrin expression. Although our studies confirm important biological effects of alphaVbeta3 integrin on enhancing cell adhesion and migration, ECM remodeling and tumor growth, beta3 integrin caused reduced MMP-2 activation in response to Col I in vitro, which appears to be physiologically relevant, as it was also seen in tumor xenografts in vivo. The reduction of MMP-2 activation (and thus MT1-MMP activity) by alphaVbeta3 in response to Col I may be important in scenarios where cells which are activated for matrix degradation need to preserve some pericellular collagen, perhaps as a substrate for cell adhesion and migration, thus maintaining a balanced level of proteolysis required for efficient tumor growth.     

Membrane type-1 matrix metalloproteinase (MT1-MMP) protects malignant cells from tumoricidal activity of re-engineered anthrax lethal toxin

Protective antigen (PA) and lethal factor (LF) are the two components of anthrax lethal toxin. PA is responsible for interacting with cell receptors and for the subsequent translocation of LF inside the cell compartment. A re-engineered toxin comprised of PA and a fusion chimera LF/Pseudomonas exotoxin (FP59) is a promising choice for tumor cell surface targeting. We demonstrated, however, that in vitro in cell-free system and in cultured human colon carcinoma LoVo, fibrosarcoma HT1080 and glioma U251 cells membrane type-1 matrix metalloproteinase (MT1-MMP) cleaves both the PA83 precursor and the PA63 mature protein. Exhaustive MT1-MMP cleavage of PA83 in vitro generates several major degradation fragments with an N-terminus at Glu40, Leu48, and Gln512. In cultured cells, MT1-MMP-dependent cleavage releases the cell-bound PA83 and PA63 species from the cell surface. As a result, MT1-MMP expressing cells have less PA63 to internalize. In agreement, our observations demonstrate that MT1-MMP proteolysis of PA makes the MT1-MMP-expressing aggressive invasive cells resistant to the cytotoxic effect of a bipartite PA/FP59 toxin. We infer from our studies that synthetic inhibitors of MMPs are likely to increase the therapeutic anti-cancer effect of anthrax toxin. In addition, our study supports a unique role of furin in the activation of PA, thereby suggesting that furin inhibitors are the likely specific drugs for short-term therapy of anthrax infection.     

Collagen binding properties of the membrane type-1 matrix metalloproteinase (MT1-MMP) hemopexin C domain. The ectodomain of the 44-kDa autocatalytic product of MT1-MMP inhibits cell invasion by disrupting native type I collagen cleavage

Up-regulation of the collagenolytic membrane type-1 matrix metalloproteinase (MT1-MMP) leads to increased MMP2 (gelatinase A) activation and MT1-MMP autolysis. The autocatalytic degradation product is a cell surface 44-kDa fragment of MT1-MMP (Gly(285)-Val(582)) in which the ectodomain consists of only the linker, hemopexin C domain and the stalk segment found before the transmembrane sequence. In the collagenases, hemopexin C domain exosites bind native collagen, which is required for triple helicase activity during collagen cleavage. Here we investigated the collagen binding properties and the role of the hemopexin C domain of MT1-MMP and of the 44-kDa MT1-MMP ectodomain in collagenolysis. Recombinant proteins, MT1-LCD (Gly(285)-Cys(508)), consisting of the linker and the hemopexin C domain, and MT1-CD (Gly(315)-Cys(508)), which consists of the hemopexin C domain only, were found to bind native type I collagen but not gelatin. Functionally, MT1-LCD inhibited collagen-induced MMP2 activation in fibroblasts, suggesting that interactions between collagen and endogenous MT1-MMP directly stimulate the cellular activation of pro-MMP2. MT1-LCD, but not MT1-CD, also blocked the cleavage of native type I collagen by MT1-MMP in vitro, indicating an important role for the MT1-MMP linker region in triple helicase activity. Similarly, soluble MT1-LCD, but not MT1-CD or peptide analogs of the MT1-MMP linker, reduced the invasion of type I collagen matrices by MDA-MB-231 cells as did the expression of recombinant 44-kDa MT1-MMP on the cell surface. Together, these studies demonstrate that generation of the 44-kDa MT1-MMP autolysis product regulates collagenolytic activity and subsequent invasive potential, suggesting a novel feedback mechanism for the control of pericellular proteolysis.     

Membrane type-1 matrix metalloproteinase (MT1-MMP) processing of pro-alphav integrin regulates cross-talk between alphavbeta3 and alpha2beta1 integrins in breast carcinoma cells

We have recently demonstrated that in breast carcinoma MCF7 cells MT1-MMP processes the alphav, alpha3, and alpha5 integrin precursors generating the respective mature S-S-linked heavy and light alpha-chains. The precursor of alpha2 integrin subunit was found resistant to MT1-MMP proteolysis. The processing of the alphav subunit by MT1-MMP facilitated alphavbeta3-dependent adhesion, activation of FAK signaling pathway, and migration of MCF7 cells on vitronectin. To elucidate further the effects of MT1-MMP on cellular integrins, we examined the functional activity of alpha5beta1 and alpha2beta1 integrins in MCF7 cells expressing MT1-MMP. Either expression of MT1-MMP alone or its coexpression with alphavbeta3 failed to affect the functionality of alpha5beta1 integrin, and adhesion of cells to fibronectin. MT1-MMP, however, profoundly affected the cross-talk involving alphavbeta3 and alpha2beta1 integrins. In MT1-MMP-deficient cells, integrin alphavbeta3 suppressed the functional activity of the collagen-binding alpha2beta1 integrin receptor and diminished cell adhesion to type I collagen. Coexpression of MT1-MMP with integrin alphavbeta3 restored the functionality of alpha2beta1 integrin and, consequently, the ability of MCF7 cells to adhere efficiently to collagen. We conclude that the MT1-MMP-controlled cross-talk between alphavbeta3 and alpha2beta1 integrins supports binding of aggressive, MT1-MMP-, and alphavbeta3 integrin-expressing malignant cells on type I collagen, the most common substratum of the extracellular matrix.     

Tissue inhibitor of metalloproteinases-2 binding to membrane-type 1 matrix metalloproteinase induces MAPK activation and cell growth by a non-proteolytic mechanism

Membrane-type 1 matrix metalloproteinase (MT1-MMP), a transmembrane proteinase with a short cytoplasmic domain and an extracellular catalytic domain, controls a variety of physiological and pathological processes through the proteolytic degradation of extracellular or transmembrane proteins. MT1-MMP forms a complex on the cell membrane with its physiological protein inhibitor, tissue inhibitor of metalloproteinases-2 (TIMP-2). Here we show that, in addition to extracellular proteolysis, MT1-MMP and TIMP-2 control cell proliferation and migration through a non-proteolytic mechanism. TIMP-2 binding to MT1-MMP induces activation of ERK1/2 by a mechanism that does not require the proteolytic activity and is mediated by the cytoplasmic tail of MT1-MMP. MT1-MMP-mediated activation of ERK1/2 up-regulates cell migration and proliferation in vitro independently of extracellular matrix proteolysis. Proteolytically inactive MT1-MMP promotes tumor growth in vivo, whereas proteolytically active MT1-MMP devoid of cytoplasmic tail does not have this effect. These findings illustrate a novel role for MT1-MMP-TIMP-2 interaction, which controls cell functions by a mechanism independent of extracellular matrix degradation.     

Membrane type 1 matrix metalloproteinase (MT1-MMP) ubiquitination at Lys581 increases cellular invasion through type I collagen

Membrane type 1 matrix metalloproteinase (MT1-MMP/MMP14) is a zinc-dependent type I transmembrane metalloproteinase playing pivotal roles in the regulation of pericellular proteolysis and cellular migration. Elevated expression levels of MT1-MMP have been demonstrated to correlate with a poor prognosis in cancer. MT1-MMP has a short intracellular domain (ICD) that has been shown to play important roles in cellular migration and invasion, although these ICD-mediated mechanisms remain poorly understood. In this study, we report that MT1-MMP is mono-ubiquitinated at its unique lysine residue (Lys(581)) within the ICD. Our data suggest that this post-translational modification is involved in MT1-MMP trafficking as well as in modulating cellular invasion through type I collagen matrices. By using an MT1-MMP Y573A mutant or the Src family inhibitor PP2, we observed that the previously described Src-dependent MT1-MMP phosphorylation is a prerequisite for ubiquitination. Taken together, these findings show for the first time an additional post-translational modification of MT1-MMP that regulates its trafficking and cellular invasion, which further emphasizes the key role of the MT1-MMP ICD.     

Membrane type 1 matrix metalloproteinase induces epithelial-to-mesenchymal transition in prostate cancer

By mining DNA microarray data bases at GenBank, we identified up-regulation of membrane type 1 matrix metalloproteinase (MT1-MMP) in human primary and metastatic prostate cancer specimens as compared with nonmalignant prostate tissues. To explore the role of up-regulated MT1-MMP in early stage cancer progression, we have employed a three-dimensional cell culture model. Minimally invasive human prostate cancer cells (LNCaP) were transfected with MT1-green fluorescent protein (GFP) chimeric cDNA as compared with GFP cDNA, and morphologic and phenotypic changes were characterized. GFP-expressing LNCaP cells formed multicellular spheroids with cuboidal-like epithelial morphology, whereas MT1-GFP-expressing cells displayed a fibroblast-like morphology and a scattered growth pattern in type I collagen gels. Cell morphologic changes were accompanied by decreased epithelial markers and enhanced mesenchymal markers, consistent with epithelial-to-mesenchymal transition. MT1-MMP-induced morphologic change and cell scattering were abrogated by target inhibition of either the catalytic domain or the hemopexin domain. We further demonstrated that MT1-MMP-induced phenotypic changes were dependent upon up-regulation of Wnt5a, which has been implicated in epithelial-to-mesenchymal transition. We conclude that MT1-MMP plays an important role in early cancer dissemination by converting epithelial cells to migratory mesenchymal-like cells.     

Membrane type I matrix metalloproteinase usurps tumor growth control imposed by the three-dimensional extracellular matrix

Cancer cells are able to proliferate at accelerated rates within the confines of a three-dimensional (3D) extracellular matrix (ECM) that is rich in type I collagen. The mechanisms used by tumor cells to circumvent endogenous antigrowth signals have yet to be clearly defined. We find that the matrix metalloproteinase, MT1-MMP, confers tumor cells with a distinct 3D growth advantage in vitro and in vivo. The replicative advantage conferred by MT1-MMP requires pericellular proteolysis of the ECM, as proliferation is fully suppressed when tumor cells are suspended in 3D gels of protease-resistant collagen. In the absence of proteolysis, tumor cells embedded in physiologically relevant ECM matrices are trapped in a compact, spherical configuration and unable to undergo changes in cell shape or cytoskeletal reorganization required for 3D growth. These observations identify MT1-MMP as a tumor-derived growth factor that regulates proliferation by controlling cell geometry within the confines of the 3D ECM.     

The role of MT2-MMP in cancer progression

The role of MT2-MMP in cancer progression remains to be elucidated in spite of many reports on MT1-MMP. Using a human fibrosarcoma cell, HT1080 and a human gastric cancer cell, TMK-1, endogenous expression of MT1-MMP or MT2-MMP was suppressed by siRNA induction to examine the influence of cancer progression in vitro and in vivo. In HT1080 cells, positive both in MT1-MMP and MT2-MMP, the migration as well as the invasion was impaired by MT1-MMP or MT2-MMP suppression. Also cell proliferation in three dimensional (3D) condition was inhibited by MT1-MMP or MT2-MMP suppression and tumor growth in the nude mice transplanted with tumor cells were reduced either MT1-MMP or MT2-MMP suppression with a prolongation of survival time in vivo. MT2-MMP suppression induces more inhibitory effects on 3D proliferation and in vivo tumor growth than MT1-MMP. On the other hand, TMK-1 cells, negative in MT1-MMP and MMP-2 but positive in MT2-MMP, all the migratory, invasive, and 3D proliferative activities in TMK-1 are decreased only by MT2-MMP suppression. These results indicate MT2-MMP might be involved in the cancer progression more than or equal to MT1-MMP independently of MMP-2 and MT1-MMP.     

Tetraspanin Proteins Regulate Membrane Type-1 Matrix Metalloproteinase-dependent Pericellular Proteolysis

Membrane type-1 matrix metalloproteinase (MT1-MMP) supports tumor cell invasion through extracellular matrix barriers containing fibrin, collagen, fibronectin, and other proteins. Here, we show that simultaneous knockdown of two or three members of the tetraspanin family (CD9, CD81, and TSPAN12) markedly decreases MT1-MMP proteolytic functions in cancer cells. Affected functions include fibronectin proteolysis, invasion and growth in three-dimensional fibrin and collagen gels, and MMP-2 activation. Tetraspanin proteins (CD9, CD81, and TSPAN2) selectively coimmunoprecipitate and colocalize with MT1-MMP. Although tetraspanins do not affect the initial biosynthesis of MT1-MMP, they do protect the newly synthesized protein from lysosomal degradation and support its delivery to the cell surface. Interfering with MT1-MMP-tetraspanin collaboration may be a useful therapeutic approach to limit cancer cell invasion and metastasis.     

Conversion of stationary to invasive tumor initiating cells (TICs): role of hypoxia in membrane type 1-matrix metalloproteinase (MT1-MMP) trafficking

Emerging evidence has implicated the role of tumor initiating cells (TICs) in the process of cancer metastasis. The mechanism underlying the conversion of TICs from stationary to invasive remains to be characterized. In this report, we employed less invasive breast cancer TICs, SK-3rd, that displays CD44(high)/CD24(low) with high mammosphere-forming and tumorigenic capacities, to investigate the mechanism by which stationary TICs are converted to invasive TICs. Invasive ability of SK-3rd TICs was markedly enhanced when the cells were cultured under hypoxic conditions. Given the role of membrane type 1-matrix metalloproteinase (MT1-MMP) in cancer invasion/metastasis, we explored a possible involvement of MT1-MMP in hypoxia-induced TIC invasion. Silencing of MT1-MMP by a shRNA approach resulted in diminution of hypoxia-induced cell invasion in vitro and metastasis in vivo. Under hypoxic conditions, MT1-MMP redistributed from cytoplasmic storage pools to the cell surface of TICs, which coincides with the increased cell invasion. In addition, CD44, a cancer stem-like cell marker, inversely correlated with increased cell surface MT1-MMP. Interestingly, cell surface MT1-MMP gradually disappeared when the hypoxia-treated cells were switched to normoxia, suggesting the plasticity of TICs in response to oxygen content. Furthermore, we dissected the pathways leading to upregulated MT1-MMP in cytoplasmic storage pools under normoxic conditions, by demonstrating a cascade involving Twist1-miR10b-HoxD10 leading to enhanced MT1-MMP expression in SK-3rd TICs. These observations suggest that MT1-MMP is a key molecule capable of executing conversion of stationary TICs to invasive TICs under hypoxic conditions and thereby controlling metastasis.     

MT1-MMP: a potent modifier of pericellular microenvironment

Cells are regulated by many different means, and there is more and more evidence emerging that changes in the microenvironment greatly affect cell function. MT1-MMP is a type I transmembrane proteinase which participates in pericellular proteolysis of extracellular matrix (ECM) macromolecules. The enzyme is cellular collagenase essential for skeletal development, cancer invasion, growth, and angiogenesis. MT1-MMP promotes cell invasion and motility by pericellular ECM degradation, shedding of CD44 and syndecan1, and by activating ERK. Thus MT1-MMP is one of the factors that influence the cellular microenvironment and thereby affect cell-signaling pathways and eventually alters cellular behavior. As a proteinase, MT1-MMP is regulated by inhibitors, but it also requires formation of a homo-oligomer complex, localization to migration front of the cells, and internalization to become a "functionally active" cell function modifier. Developing new means to inhibit "functional activity" of MT1-MMP may be a new direction to establish treatments for the diseases that MT1-MMP mediates such as cancer and rheumatoid arthritis.     

Inhibition of membrane type-1 matrix metalloproteinase by cancer drugs interferes with the homing of diabetogenic T cells into the pancreas

We have discovered that clinically tested inhibitors of matrix metalloproteinases can control the functional activity of T cell membrane type-1 matrix metalloproteinase (MT1-MMP) and the onset of disease in a rodent model of type 1 diabetes in non-obese diabetic mice. We determined that MT1-MMP proteolysis of the T cell surface CD44 adhesion receptor affects the homing of T cells into the pancreas. We also determined that both the induction of the intrinsic T cell MT1-MMP activity and the shedding of cellular CD44 follow the adhesion of insulin-specific, CD8-positive, Kd-restricted T cells to the matrix. Conversely, inhibition of these events by AG3340 (a potent hydroxamate inhibitor that was widely used in clinical trials in cancer patents) impedes the transmigration of diabetogenic T cells into the pancreas and protects non-obese diabetic mice from diabetes onset. Overall, our studies have divulged a previously unknown function of MT1-MMP and identified a promising novel drug target in type I diabetes.     

Oxidative stress and prostate cancer progression are elicited by membrane-type 1 matrix metalloproteinase

Oxidative stress caused by high levels of reactive oxygen species (ROS) has been correlated with prostate cancer aggressiveness. Expression of membrane-type 1 matrix metalloproteinase (MT1-MMP), which has been implicated in cancer invasion and metastasis, is associated with advanced prostate cancer. We show here that MT1-MMP plays a key role in eliciting oxidative stress in prostate cancer cells. Stable MT1-MMP expression in less invasive LNCaP prostate cancer cells with low endogenous MT1-MMP increased activity of ROS, whereas MT1-MMP knockdown in DU145 cells with high endogenous MT1-MMP decreased activity of ROS. Expression of MT1-MMP increased oxidative DNA damage in LNCaP and in DU145 cells, indicating that MT1-MMP-mediated induction of ROS caused oxidative stress. MT1-MMP expression promoted a more aggressive phenotype in LNCaP cells that was dependent on elaboration of ROS. Blocking ROS activity using the ROS scavenger N-acetylcysteine abrogated MT1-MMP-mediated increase in cell migration and invasion. MT1-MMP-expressing LNCaP cells displayed an enhanced ability to grow in soft agar that required increased ROS. Using cells expressing MT1-MMP mutant cDNAs, we showed that ROS activation entails cell surface MT1-MMP proteolytic activity. Induction of ROS in prostate cancer cells expressing MT1-MMP required adhesion to extracellular matrix proteins and was impeded by anti-β1 integrin antibodies. These results highlight a novel mechanism of malignant progression in prostate cancer cells that involves β1 integrin-mediated adhesion, in concert with MT1-MMP proteolytic activity, to elicit oxidative stress and induction of a more invasive phenotype.