Steroid hormones receptor characterization

The receptors of steroid hormones were the first representatives of the family of nuclear receptors to be characterized. With the characterization of further receptors it became clear that the signaling pathway of the nuclear receptors differ significantly in detail. Based on the receptor activation mechanism the nuclear receptors can be divided into two basic groups ... 

Vitamin D3 is a precursor of the hormone 1,25-dihy-droxyvitamin D3. Vitamin D3 is essential for normal calcium and phosphorus metabolism. It is formed from 7-dehydrocholesterol by ultraviolet photolysis in the skin. Insufficient exposure to sunlight and absence of vitamin D3 in the diet leads to rickets, a condition characterized by weak, malformed bones. Vitamin D3 is inactive, but it is converted into an active compound by two hydroxylation reactions that occur in different organs. The first hydroxylation occurs in the liver, which produces 25-hydroxyvita-min D3, abbreviated 25(OH)D3 the second hydroxylation occurs in the kidney and gives rise to the active product 1,25-dihydroxy vitamin D3 24,25 (OH)2D3 (fig. 24.13). The hydroxylation at position 1 that occurs in the kidney is stimulated by parathyroid hormone (PTH), which is secreted from the parathyroid gland in response to low circulating levels of calcium. In the presence of adequate calcium, 25(OH)D3 is converted into an inactive metabolite, 24,25 (OH)2D3. The active derivative of vitamin D3 is considered a hormone because it is transported from the kidneys to target cells, where it binds to nuclear receptors that are analogous to those of typical steroid hormones. l,25(OH)2D3 stimulates calcium transport by intestinal cells and increases calcium uptake by osteoblasts (precursors of bone cells). 

Monkeys, in particular Rhesus macaque and squirrel monkey, have been considered as models for vaginal drug absorption because of the similarities between nonhuman primate and human anatomy. The vagina of rhesus macaques is characterized by connective tissue attachments, localization of steroid hormone receptors, duration, and cyclic changes in vaginal physiology similar to that of humans [119]. 

Steroid hormones achieve their effects on target tissues through intracellular receptor proteins. According to recent views, oestrogen and progestin receptors are localized in the nuclear compartment of the cells, whereas glucocorticoid receptors may reside in both the cytoplasm and the nucleus. Determination of the intracellular localization of androgen receptors awaits the development of (monoclonal) antibodies which will enable immunohistochemical studies. The molecular aspects of the mechanism of action of steroid hormones will be covered in other chapters [1-3] in this volume. The present chapter deals with the characterization, assay and purification of steroid receptors. 

There have been a number of technological advances in recent years which are responsible for much of the progress made in our understanding of steroid receptor mechanisms. These include the ability to purify receptors, affinity labeling techniques, production of antireceptor antibodies for direct detection of receptor proteins, and isolation and characterization of hormone responsive genes as well as the genes for receptors themselves. 

The l,25(OH)2D3 receptor of the chick intestine has been extensively characterized by our laboratory and several others, and its biochemical properties reveal many similarities to classical steroid hormone receptors [5,6], The l,25(OH)2D3 receptor is a protein with a molecular mass of approximately 67000 Da when occupied, unoccupied or in the absence of protease inhibitors. The receptor has a high affinity for l,25(OH)2D3 with a Kd in the range of 1-50 x 10 n M. The specificity of the receptor for binding of l,25(OH)2D3 metabolites and analogues, as determined by competitive binding studies, parallels the biological activity of these compounds. 

S. A. Ofiate, S. Y. Tsai, M.-J. Tsay, B. W. O Malley. Sequence and characterization of a coactivator for the steroid hormone receptor superfamily. Science, 270, 1354r-1357, 1995. 

Miesfeld, R., Okret, S., Wikstrom, A.C., Wrange, O., Gustafsson, J.A. and Yamamoto, K.R. (1984) Characterization of a steroid hormone receptor gene and mRNA in wild-type and mutant cells. Nature, 312, 779-781. 

Like vertebrate steroid hormones, ecdysteroids recognize target tissues by binding with nuclear receptors. The ecdysteroid receptor has been isolated and characterized from the fiddler crab, Uca pugilator. It has been sequenced and has homologies with insect ecdysteroid receptors. Transcripts for the receptor were isolated from crab limb buds and developing ovaries. 

A study of the relationship between dietary factors and breast tumor characteristics was carried out in premenopausal women (Touillaud et al., 2005). Tumors were characterized in terms of their estrogen receptor (ER) status. Tumors with ER-positive status, as compared to ER-negative, are known to respond to the proliferative effect of estrogen, and are more likely to react to endocrine therapy. In the study, a low intake of boron (0.8 mg/day) was associated with a lower risk of ER-negative tumors as compared to a higher intake (1.03 mg/day). A similar relationship was observed with phytochemicals, such as phytosterols and kaempferol. Although these data are preliminary, they suggest that the effect of boron is confounded by the presence of phytosterols that are known to interfere with the entero-hepatic circulation of steroids, steroid hormones, and cholesterol (Samman et al, 2004). 

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