The goal of this project is the characterization of the mechanism(s) by which thyroid hormone regulates actin polymerization and, thus, affects the integrity of the actin cytoskeleton in cells of the Central Nervous System. Because alterations in polymerization of the actin cytoskeleton would significantly affects the complex interactions between the cytoskeletal network and the extracellular matrix, resulting in attenuation of neuronal growth and development, studies of the interactions of T4 with the actin cytoskeleton will provide insight into thyroid hormone's role in the growth and differentiation of neurons. The model system to be examined is the T4- dependent regulation of type II iodothyronine 5'-deiodinase (5'D-II) in cultured, cAMP-stimulated glial cells. T4 modulates levels of this important cerebrocortical enzyme by decreasing the biological half-life of 5'D-II through a novel, extra-nuclear mode of action. Disruption of the microfilaments blocks this T4-mediated action, indicating that the actin cytoskeleton is a likely site for regulation by thyroid hormone. 5'D-II can be affinity labeled by a T4 derivative and, thus, can be visualized in the cell using immunocytochemical and histochemical techniques. The morphological characterization of the interactions between T4, actin and 5'D-II will be accomplished during the first phase of this project. The second phase of the project will involve the identification of the mediator(s) of thyroid hormone's effects on the cytoskeleton. Actin and T4 affinity chromatography will be used to isolate potential actin- binding proteins which will then be evaluated in a cell free system to asses their ability to affect actin polymerization. Protein(s) that bind to filaments actin and T4 and that demonstrate a T4-modulated effect on actin polymerization will be characterized further with 2D SDS-PAGE and peptide mapping and compared to information available on actin-binding proteins previously described. The identification and characterization of the mechanism(s) by which T4 interacts with the cytoskeleton, and thereby regulates the turnover of a short-lived membrane protein, will then allow the study of the more fundamental questions regarding thyroid hormone's influence on, and participation in, the growth of developing neurons and the subsequent establishment of cell-cell and cell-substratum attachments.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08DK002005-03
Application #
3080923
Study Section
Diabetes, Endocrinology and Metabolic Diseases B Subcommittee (DDK)
Project Start
1990-07-01
Project End
1995-06-30
Budget Start
1992-07-01
Budget End
1993-06-30
Support Year
3
Fiscal Year
1992
Total Cost
Indirect Cost
Name
University of Massachusetts Medical School Worcester
Department
Type
Schools of Medicine
DUNS #
660735098
City
Worcester
State
MA
Country
United States
Zip Code
01655
Leonard, J L; Leonard, D M; Safran, M et al. (1999) The mammalian homolog of the frog type II selenodeiodinase does not encode a functional enzyme in the rat. Endocrinology 140:2206-15
Leonard, J L; Farwell, A P (1997) Thyroid hormone-regulated actin polymerization in brain. Thyroid 7:147-51
Safran, M; Farwell, A P; Leonard, J L (1996) Catalytic activity of type II iodothyronine 5'-deiodinase polypeptide is dependent upon a cyclic AMP activation factor. J Biol Chem 271:16363-8
Leonard, J L; Leonard, D M; Shen, Q et al. (1996) Selenium-regulated translation control of heterologous gene expression: normal function of selenocysteine-substituted gene products. J Cell Biochem 61:410-9
Farwell, A P; Safran, M; Dubord, S et al. (1996) Degradation and recycling of the substrate-binding subunit of type II iodothyronine 5'-deiodinase in astrocytes. J Biol Chem 271:16369-74
Farwell, A P; Tranter, M P; Leonard, J L (1995) Thyroxine-dependent regulation of integrin-laminin interactions in astrocytes. Endocrinology 136:3909-15
Leonard, J L; Farwell, A P; Yen, P M et al. (1994) Differential expression of thyroid hormone receptor isoforms in neurons and astroglial cells. Endocrinology 135:548-55
Farwell, A P; DiBenedetto, D J; Leonard, J L (1993) Thyroxine targets different pathways of internalization of type II iodothyronine 5'-deiodinase in astrocytes. J Biol Chem 268:5055-62
Safran, M; Farwell, A P; Rokos, H et al. (1993) Structural requirements of iodothyronines for the rapid inactivation and internalization of type II iodothyronine 5'-deiodinase in glial cells. J Biol Chem 268:14224-9
Chanoine, J P; Safran, M; Farwell, A P et al. (1992) Effects of selenium deficiency on thyroid hormone economy in rats. Endocrinology 131:1787-92

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