The aim of our project remains to understand effects of glucocorticoid hormones on gene expression. We are investigating two responses: the lysis of T-lymphoma cells and the induction of fibronectin in a fibrosarcoma cell line. Our studies of glucocorticoid-induced lymphocytolysis have been done so far with murine lymphoid cell lines. In recent months we have extended our work to the human acute lymphoblastic leukemic cell line CEM. We have examined the basis for glucocorticoid resistance in a subclone of that line, CEM-C1, known to contain functional glucocorticoid receptor. Treatment of CEM-C1 cells with the DNA-demethylating agent 5-aza-2' deoxycytidine yielded glucocorticoid-sensitive subclones at high frequency (approximately 10%). This indicates that DNA methylation is responsible for inactivating the expression of a """"""""lytic"""""""" function required for glucocorticoid-sensitivity in these cells. We have followed up on our observation that glucocorticoids increase 10- to 20-fold the rate of fibronectin biosynthesis in HT1080 fibrosarcoma cells. Using a human fibronectin cDNA clone to probe Northern blots, we found that this probe hybridizes with a high molecular weight mRNA from dexamethasone-treated HT1080. This mRNA is present at a much lower level in untreated HT1080 cells and is undetectable in CEM cells, which do not produce fibronectin. By """"""""slot"""""""" blot analysis of total cellular RNA, we found a 16- to 32-fold increase in fibronectin RNA level after hormone treatment. Moreover, this induction is inhibited by a glucocorticoid antagonist, RU486. This study demonstrates that the dexamethasone-induced stimulation of fibronectin biosynthesis is the result of an increase in accumulated fibronectin mRNA mediated by the glucocorticoid receptor. Studies are currently in progress to examine whether this effect is due to an increased rate of transcription. (D)

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA036146-14
Application #
3173653
Study Section
Mammalian Genetics Study Section (MGN)
Project Start
1978-12-01
Project End
1988-11-30
Budget Start
1986-12-01
Budget End
1987-11-30
Support Year
14
Fiscal Year
1987
Total Cost
Indirect Cost
Name
Salk Institute for Biological Studies
Department
Type
DUNS #
005436803
City
La Jolla
State
CA
Country
United States
Zip Code
92037
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Baughman, G; Harrigan, M T; Campbell, N F et al. (1991) Genes newly identified as regulated by glucocorticoids in murine thymocytes. Mol Endocrinol 5:637-44
Harrigan, M T; Baughman, G; Campbell, N F et al. (1989) Isolation and characterization of glucocorticoid- and cyclic AMP-induced genes in T lymphocytes. Mol Cell Biol 9:3438-46
Gruol, D J; Rajah, F M; Bourgeois, S (1989) Cyclic AMP-dependent protein kinase modulation of the glucocorticoid-induced cytolytic response in murine T-lymphoma cells. Mol Endocrinol 3:2119-27
Dean, D C; Newby, R F; Bourgeois, S (1988) Regulation of fibronectin biosynthesis by dexamethasone, transforming growth factor beta, and cAMP in human cell lines. J Cell Biol 106:2159-70

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