Human thrombospondin (TSP) is a high molecular weight glycoprotein composed of three identical disulphide bonded peptide chains. Originally identified as a platelet alpha-granule protein essential for the secondary phase of platelet aggregation, TSP has now been shown to be synthesized by many different cell types including epithelial cells, fibroblasts, smooth muscle cells, endothelial cells and monocytes. In these cells the synthesized TSP is secreted and incorporated into the extracellular matrix (ECM). We have recently shown that TSP is synthesized, secreted and bound to the cell surface by both human keratinocytes and squamous carcinoma cells where it mediates cell-cell and cell-substrate interactions. Further, there is an inverse relationship between the state of differentiation and the ability to synthesize and utilize TSP. Thus, cells that are least differentiated produce the greatest amount of TSP and have the highest number of cell surface receptors for it. We find that there are two classes of receptor for TSP on squamous carcinoma cells that recognize and bind to different regions of the TSP molecule. There is a moderate affinity/high number class of receptor that binds to the heparin binding domain of TSP. This receptor has the characteristics of a heparan sulphate proteoglycan. The other class of receptors is high affinity/low number and binds to the 140 kD fragment of TSP that does not contain the heparin binding domain. This receptor is composed of two cell surface proteins (150/80 kD) and mediates the attachment of squamous carcinoma cells to TSP coated dishes (cell- substrate interaction). In order to better understand the structure and functional role of the 105/80 kD cell adhesion promoting receptor, we propose the following.
Specific Aim 1 : Examine the contribution of the 105/80 kD TSP binding cell surface proteins in cell attachment and cell differentiation. Studies will involve cell attachment and radioligand binding assays, generation of polyclonal and monoclonal antibodies and protein microsequencing of the receptor proteins.
Specific Aim 2 : cDNA cloning of the TSP-Receptor(s). This will be accomplished by screening squamous cell cDNA expression libraries with a combination of antibody and oligonucleotide probes. The cloned cDNA's will be used to study the regulation of the TSP receptor(s) during differentiation.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA051888-03
Application #
3196605
Study Section
Pathobiochemistry Study Section (PBC)
Project Start
1991-02-05
Project End
1994-01-31
Budget Start
1993-03-15
Budget End
1994-01-31
Support Year
3
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Type
Schools of Medicine
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109