Evidence for a direct interaction between the tumor suppressor serpin, maspin, and types I and III collagen.
Maspin (mammary serine protease inhibitor) was originally identified as a tumor suppressor protein in human breast epithelial cells and is a member of the serine proteases inhibitor (serpin) superfamily. It inhibits tumor cell motility and angiogenesis, and although predominantly cytoplasmic, it is also localized to the cell surface. In this study we have investigated the use of the yeast two-hybrid interaction trap to identify novel maspin targets. A target human fibroblast cDNA library was screened, and the alpha-2 chain of type I collagen was identified as a potential interactant. Binding studies with isolated proteins showed interaction between recombinant maspin and types I and III collagen but not other collagen subtypes, a profile strikingly similar to mouse pigment epithelium-derived factor (caspin), which is similarly down-regulated in murine adenocarcinoma tumors and is a potent inhibitor of angiogenesis. Kinetic analysis using an IAsys resonant mirror biosensor determined the dissociation constant of maspin for collagen type I to be 0.63 microm. Further two-hybrid interactions with maspin truncation constructs suggest that collagen binding is localized to amino acids 84-112 of maspin, which aligns with the collagen-binding region of colligin. A direct interaction between exogenous or cell surface maspin and extracellular matrix collagen may contribute to a cell adhesion role in the prevention of tumor cell migration and angiogenesis.
Blacque
Oliver E
OE
Department of Biochemistry and Conway Institute for Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
Worrall
D Margaret
DM
D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D024042:Collagen Type I / Q000378:metabolism*; D024061:Collagen Type III / Q000378:metabolism*; D017931:DNA Primers; D016147:Genes, Tumor Suppressor; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D016133:Polymerase Chain Reaction; D011506:Proteins / Q000378:metabolism*; D015843:Serpins / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D024042:Collagen Type I; D024061:Collagen Type III; D017931:DNA Primers; D011506:Proteins; C085344:SERPIN-B5; D015843:Serpins