Degree Type

Dissertation

Date of Award

1990

Degree Name

Doctor of Philosophy

Department

Biochemistry, Biophysics and Molecular Biology

First Advisor

Marit Nilsen-Hamilton

Abstract

A series of studies were conducted to purify, characterize, sequence, and evaluate the mechanism of expression of a secreted, superinducible protein, SIP 24. It has been found that the potent tumor promoter, 12-o-tetradecanoyl-13-acetate (TPA) in combination with the protein synthesis inhibitor, cycloheximide (CHX), can synergistically increase SIP 24 expression. The in vivo activator of protein kinase C (PKC), diacylglycerol, also increased SIP 24 expression. Further evidence that PKC is involved in regulating SIP 24 expression, was found when an inhibitor of PKC, H7, was added to cells stimulated by TPA and bFGF, and SIP 24 levels were reduced. Studies, in which PKC was down-regulated by saturation with TPA, also indicate that both TPA and basic fibroblast growth factor (bFGF) act through the protein kinase C pathway to increase SIP 24 production;The glucocorticoid analog, dexamethasone, acted synergistically with CHX to increase SIP 24 expression. Agents, which increase intracellular cAMP levels, increased SIP 24 expression, but it is not clear whether the involvement of the cAMP-mediated pathway is of physiological importance to SIP 24 expression;SIP 24 was purified to homogeneity, polyclonal antibodies were obtained, and a radioimmunoassay was developed to quantitate SIP 24. SIP 24 is a glycoprotein with 3-4,000 daltons of N-linked carbohydrate. By using reducing versus non-reducing SDS-polyacrylamide electrophoresis, we determined that SIP 24 has intramolecular disulfide bonds. A partial amino acid sequence of SIP 24 was determined from analysis of peptides derived by clostripain digestion. Two of the peptides were found to have a high sequence identity to bovine cyclophilin, a cyclosporin A binding protein. If we find that SIP 24 can bind the potent immunosuppressive agent, cyclosporin A, this would establish a link between growth factors and regulation of an immune response.

DOI

https://doi.org/10.31274/rtd-180813-9168

Publisher

Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/

Copyright Owner

Thomas Richard Davis

Language

en

Proquest ID

AAI9101346

File Format

application/pdf

File Size

92 pages

Included in

Biochemistry Commons

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