Some Properties of the Ecdysteroid Receptor in the Salivary Gland of the Ixodid Tick, Amblyomma hebraeum

Abstract

Salivary gland degeneration in ixodid ticks is triggered by an ecdysteroid hormone. We used [3H]ponasterone A (PoA) as a specific ligand to detect the ecdysteroid receptor in the salivary glands of large partially fed female ticks (Amblyomma hebraeum Koch; Acari; Ixodidae). Binding of [3H]PoA was thermolabile and sensitive to pronase, but not to DNase or RNase, indicating that the ligand binds to a protein. Scatchard analysis of [3H]PoA binding strongly suggested the presence of an ecdysteroid receptor in cytosolic and nuclear extracts of the tissue. The Kd and Bmax for PoA binding in cytosol were 0.72 ± 0.09 nM and 175 ± 12 fmol/mg protein, respective (n = 8). Corresponding figures for neclear extract were 1.1 ± 0.5 nM and 282 ± 35 fmol/mg protein, respectively (n = 3; P > 0.05 compared to cytosol). The relative ability of unlabeled ecdysteroids to compete for [3H]PoA binding was (in descending order): PoA > muristerone A > makisterone A > 20-hydroxyecdysone > mesylinokosterone > ecdysone. The Kd estimated for 20-hydroxyecdysone (probably the natural hormone) correlates very well with its physiological potency in inducing salivary gland degeneration in vivo and in organ culture. None of the vertebrate steroids tested (estradiol, testosterone, progesterone, and corticosterone) was able to displace PoA binding at a concentration 105 times higher than PoA. The cytosolic form of the receptor migrated to the 3.2 S region of a 10-40% sucrose density gradient.