Identification of CD63 as a tissue inhibitor of metalloproteinase-1 interacting cell surface protein

Abstract

This study identified CD63, a member of the tetraspanin family, as a TIMP-1 interacting protein by yeast twohybrid screening. Immunoprecipitation and confocal microscopic analysis confirmed CD63 interactions with TIMP-1, integrin b1, and their co-localizations on the cell surface of human breast epithelial MCF10A cells. TIMP-1 expression correlated with the level of active integrin b1 on the cell surface independent of cell adhesion. While MCF10A cells within a three-dimensional (3D) matrigel matrix form polarized acinar-like structures, TIMP-1 overexpression disrupted breast epithelial cell polarization and inhibited caspase-mediated apoptosis in centrally located cells, necessary for the formation and maintenance of the hollow acinar-like structures. Small hairpin RNA (shRNA)-mediated CD63 downregulation effectively reduced TIMP-1 binding to the cell surface, TIMP-1 colocalization with integrin b1, and consequently reversed TIMP-1-mediated integrin b1 activation, cell survival signaling and apoptosis inhibition. CD63 downregulation also restored polarization and apoptosis of TIMP-1 overexpressing MCF10A cells within a 3D-matrigel matrix. Taken together, the present study identified CD63 as a cell surface binding partner for TIMP-1, regulating cell survival and polarization via TIMP-1 modulation of tetraspanin/integrin signaling complex.

Adiponectin Reduces Hepatic Stellate Cell Migration by Promoting Tissue Inhibitor of Metalloproteinase-1 (TIMP-1) Secretion

“…This highly controversial observation in terms of current views on the role of TIMP-1 in fibrosis led us to speculate on the potential biological relationship between adiponectin, TIMP-1, and FAK during liver fibrosis. Previous studies have shown that TIMP-1 can have MMP-independent signaling through CD63 (Tspan 30), a member of the tetraspanins (23). These molecules are hydrophobic proteins containing four transmembrane ␣-helices, two extracellular loops, and a short cytoplasmic tail.…”

“…TIMP-1 through the CD63/␤1-Integrin Complex Impairs HSC Motility-It has previously been reported that TIMP-1 regulates FAK activity through MMP-independent signaling by binding to the tetraspanin CD63/␤1-integrin complex (23). We investigated whether this complex was present in HSCs.…”

The tetraspanin CD63 is involved in granule targeting of neutrophil elastase

“…21 Tetraspanin complexes act as molecular facilitators in many cellular processes, 35,36 and different partner proteins can become clustered together with CD63. 37 It has been proposed that membrane proteins such as the H,K-ATPase 25 and membrane type 1-matrix metalloproteinase 23,38 are transported by CD63. Our study suggests that soluble proteins may also be escorted by CD63.…”

Tetraspanin CD63 Promotes Vascular Endothelial Growth Factor Receptor 2-β1 Integrin Complex Formation, Thereby Regulating Activation and Downstream Signaling in Endothelial Cells in Vitro and in Vivo

“…A number of other cell types have also been shown to depend on CD63 for efficient adhesion, spread, and migration including platelets, monocytes, neutrophils, dendritic, intestinal, and melanoma cells (14 -19). In addition, CD63 has been implicated in the regulation of cell survival and polarization in breast epithelial cells, where it was found to interact with the tissue inhibitor of metalloproteinase-1 (20). Interestingly, CD63 is one of several lysosomal proteins affected in Hermansky-Pudlak syndrome with adaptor protein complex-3 deficiency, a rare autosomal recessive disorder characterized by defective intracellular vesicle formation and a platelet storage pool deficiency (21).…”