Characterization of putative steroid receptors in the membrane, cytosol and nuclear fractions from the olfactory tissue of brown and rainbow trout |
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Authors: | T G Pottinger A Moore |
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Institution: | (1) The Institute of Freshwater Ecology, Windermere Laboratory, The Ferry House, Far Sawrey, Ambleside, Cumbria, LA22 0LP, UK;(2) MAFF Fisheries Laboratory, Lowestoft, Suffolk, NR33 0HT, UK |
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Abstract: | Specific binding sites for testosterone have been detected in three compartments of olfactory tissue from brown and rainbow
trout. Binding of3H-testosterone to the membrane fraction of olfactory tissue is of high affinity (Kd = 0.5–1.9 nM) and limited capacity (Nmax = 30–60 fmol mg+1 protein). Binding is reversible, and is eliminated by protease treatment. The membrane binding site exhibits a high degree
of ligand specificity; 11β-hydroxytestosterone, 11-ketotestosterone, 17α-hydroxyprogesterone, 17α,20β-dihydroxy-4-pregnen-3-one,
cortisol, and estradiol-17β all fail to displace testosterone at 20-fold excess while testosterone itself competes successfully.
These attributes are consistent with the presence of specific steroid receptor proteins. Binding of testosterone within the
cytosol is of moderate affinity (Kd = 9.0–23.0 nM) and high capacity (Nmax = 0.5–2.9 pmol mg+1 protein) and is more readily displaced by a number of steroid competitors than is the case for the membrane site. The rate
of association and dissociation of testosterone from the cytosolic binding site is markedly more rapid than the equivalent
processes in the membrane fraction. Binding of testosterone to the nuclear extract is of high affinity (Kd ∼3.0 nM) and limited capacity (Nmax ∼50 fmol mg+1 protein).
There are no substantial differences between species or between sexes in the affinity or capacity of testosterone-binding
sites in nuclear extract or membrane fraction. However, cytosolic testosterone-binding sites are three- to four-fold more
abundant in rainbow trout than in brown trout, and female rainbow trout have more cytosolic binding sites than male rainbow
trout, but a lower affinity for testosterone than male sites.
Preliminary evidence supports the involvement of the membrane-associated testosterone-binding site in olfactory processes.
Rainbow trout display an EOG response to testosterone at a concentration (≥ 10+9 M) which is consistent with the equilibrium dissociation constant (Kd) of the membrane-associated testosterone-binding site. Binding of3H-testosterone to the membrane-associated site shows a pH dependency which is comparable to the effects of pH on the EOG response
to testosterone in intact fish. The attributes of the intracellular testosterone-binding sites are common to testosterone
receptors in other fish tissues which are known androgen target tissues. This suggests that the development and/or function
of salmonid olfactory tissue may be susceptible to influence by endogenous testosterone. |
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Keywords: | rainbow trout brown trout steroid receptors olfactory tissue pheromones |
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