IMMUNOCHEMICAL CHARACTERIZATION OF PARATHYROID HORMONE-RELATED PROTEIN FROM TUMOR AND NONTUMOUR CELLS

The molecular forms of parathyroid hormone-related protein (PTHRP) in conditioned media from the BEN human lung cancer cell line, rat parathyroid cells (PT-r) and human keratinocytes were studied by gel-filtration chromatography with assay of PTHRP by immunoassays and bioassay. Immunoreactivity (1-86 and 1-34) and bioactivity (1-34) in conditioned media eluted as a coincident major peak (approx. molecular mass 19-22 kDa) and there was evidence of amino-terminal species in the molecular mass range 10-16 kDa in BEN and keratinocyte media. Western blotting of PTHRP affinity purified by monoclonal antibodies directed at regions 1-34 or 37-67, identified a major species in all cell cytosols and media with an apparent molecular mass of 24-25 kDa, consistently slightly larger than recombinant PTHRP(1-141) (mobility of 21 kDa) which may represent an intact or native form of PTHRP. Additional amino-terminal species were identified in medium from keratinocytes (16 and 7 kDa), BEN cells (18 and 14 kDa) and PT-R cells (17 kDa), suggesting that processing occurs at the C-terminus and within the mid-region to form a range of amino-terminal fragments.