Centrosomal Pericentrin Is a Direct Cleavage Target of Membrane Type-1 Matrix Metalloproteinase in Humans but Not in Mice

Abstract

Membrane type-1 matrix metalloproteinase (MT1-MMP) exhibits distinctive and important pericellular cleavage functions. Recently, we determined that MT1-MMP was trafficked to the centrosomes in the course of endocytosis. Our data suggested that the functionally important, integral, centrosomal protein, pericentrin-2, was a cleavage target of MT1-MMP in human and in canine cells and that the sequence of the cleavage sites were ALRRLLG 1156 2L 1157 FG and ALRRLLS 2068 2L 2069 FG, respectively. The presence of Asp-948 at the P1 position inactivated the corresponding site (ALRRLLD 948 -L 949 FGD) in murine pericentrin. To confirm that MT1-MMP itself cleaves pericentrin directly, rather than indirectly, we analyzed the cleavage of the peptides that span the MT1-MMP cleavage site. In addition, we analyzed glioma U251 cells, which co-expressed MT1-MMP with the wild type murine pericentrin and the D948G mutant. We determined that the D948G mutant that exhibited the cleavage sequence of human pericentrin was sensitive to MT1-MMP, whereas unmodified murine pericentrin was resistant to proteolysis. Taken together, our results confirm that MT1-MMP cleaves pericentrin-2 in humans but not in mice and that mouse models of cancer probably cannot be used to critically examine MT1-MMP functionality. MT1-MMP2 /MMP-14 is a prototypic member of the membranetethered matrix metalloproteinases (1). Although MT1-MMP is present in normal tissues, its enhanced expression is directly linked to tumor progression and metastasis (2-6). Cell surface-associated MT1-MMP is a multifunctional enzyme (7), and it is involved in the pericellular proteolysis of the extracellular matrix, the activation of soluble MMPs, and the cleavage of adhesion and signaling cell receptors (8 -10). The functional activity of MT1-MMP is regulated by its activation by furinlike proprotein convertases, by its inhibition by TIMPs, and by its selfproteolysis and shedding (11-13). Evidence is also emerging that exocytosis, endocytosis, and recycling also regulate the presentation of MT1-MMP on the cell surface and, consequently, its cell surface-associated proteolytic activity (14 -17).Recently, we determined that functionally active MT1-MMP, which was presented on the cell surface, was internalized, trafficked alongside the microtubular cytoskeleton, and delivered to the centrosomal compartment (16,18). The presence of MT1-MMP in the pericentrosomal space correlated with the cleavage of human pericentrin-2 (kendrin), an integral and functionally important centrosomal, 3336-amino-acid residue long, protein (19 -22), and chromosome instability in non-malignant epithelial Madin-Darby canine kidney cells (18,23). Centrosomes, spindle pole bodies, are cellular organelles that exhibit an ability to organize microtubules and to nucleate (24). The normal functionality of centrosomes is essential to the organization of the cytoskeleton and the mitotic spindle, self-duplication, and cell cycle progression (25, 26). Conversely, centrosomal abnormalities, early predictors of carcinog...

Proteolysis-Driven Oncogenesis

“…MT1-MMP is present also in the Rab-11-positive vesicles in the pericentrosomal compartment. 18,34,35 In the metaphase cells, MT1-MMP is partially colocalized �ith the centrosomes. MT1-MMP extensively accumulates in the midbody at the late sta�e of cytokinesis.…”

“…In a�reement, the D948G murine pericentrin mutant that exhibited the human cleava�e site �as susceptible to MT1-MMP proteolysis. 34 Based on these data, �e concluded that mouse models of cancer, unfortunately, cannot be used to critically examine the role of MT1-MMP in incipient cancer in humans. Overall, our findin�s �arrant additional studies of the �enetically redesi�ned animal models, so that they �ill fully recapitulate human tumori�enesis.…”

“…A mass-spectrometry analysis of the di�est determined the mass of the cleava�e fra�ments and, consequently, allo�ed us to identify the scissile bond (G 672 ↓ L 673 and G 1156 ↓L 1157 ). 18,34 Endo�enous pericentrin �as efficiently cleaved in human �lioma U251, breast carcinoma MCF7 and normal mammary epithelial 184B5 cells and in canine Madin-Darby canine kidney (MDCK) cells transfected �ith MT1-MMP. In a�reement, the 200 kDa and 150 kDa de�raded forms of pericentrin �ere identified in U251 cells.…”

Membrane Type-1 Matrix Metalloproteinase Confers Aneuploidy and Tumorigenicity on Mammary Epithelial Cells

“…FG motif of pericentrin B and induces chromosomal instability and aneuploidy (22,23). The transmembrane domain is critical for the trafficking of MT1-MMP to the PCM, suggesting that internalized MT1-MMP is integrated in the vesicle membrane (24).…”

Η Επίδραση Των Μεταλλοπρωτεϊνασών Στη Διαπερατότητα Του Υπεζωκότα

“… Οι MMP2, 7, 9 και ιδιαίτερα η 14 φαίνεται ότι εκφράζονται στους περισσότερους καρκινικούς όγκους [133].…”