Date of Award


Degree Type


Degree Name

Doctor of Philosophy


The murine cDNA encoding secreted phosphoprotein 1 (SPP) was cloned in this lab on the basis of induction of the mRNA (originally designated 2ar) by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate in JB6 epidermal cells. The polypeptide predicted from the cDNA sequence is 84% identical to rat osteopontin (44 kDa bone phosphoprotein; sialoprotein I). Many primary structural features are conserved, including an ArgGlyAsp (RGD) cell adhesion site and an unusual series of 9-10 consecutive aspartate residues. SPP mRNA is most abundant in vivo in bone and kidney, and was localized to the medulla region of adult mouse kidney by in situ hybridization. Portions of the predicted SPP polypeptide were expressed as cro-{dollar}\beta{dollar}-galactosidase fusion proteins, and polyclonal antisera were obtained. The SPP antisera recognized the major ({dollar}\sp{lcub}32{rcub}{dollar}P) orthophosphate-labeled protein secreted by all mouse and rat cell lines studied. This observation led to the discovery that SPP is closely related or identical to a transformation-related phosphoprotein, also known as pp69. SPP mRNA was transiently induced by treatment of mouse epidermis in vivo with 12-O-tetradecanoylphorbol-13-acetate, and was constitutively expressed in 2 of 3 epidermal papillomas and 7 of 7 squamous cell carcinomas. SPP expression was elevated in T24 H-ras-transformed mouse fibroblasts, and the level of expression correlated with the metastatic ability of these cells. It is proposed that SPP may act as an autocrine adhesion factor for tumor cells. SPP is encoded by a single copy mouse gene which spans approximately 9 kilobasepairs (kb). A genomic clone containing the first 6 exons and 16 kb of 5{dollar}\sp\prime{dollar} flanking DNA was isolated. The SPP promoter region contains a TATA-like box (TTTAAA), a CCAAT box (reverse complement), and a number of potential regulatory elements. A small fragment ({dollar}-{dollar}235 to +79) was able to direct high level tumor promoter-inducible expression of a fused marker gene.



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