Vitellogenin gene expression in male Xenopus hepatocytes during primary and secondary stimulation with estrogen in cell cultures

Abstract

Primary cultures of male Xenopus liver parenchymal cells that retained their competence to respond to estrogen were used to study the hormone-induced activation of the vitellogenin gene in vitro. The accumulation of vitellogenin mRNA in these cells was monitored by a quantitative diazotized paper disc hybridization procedure with a sensitivity of at least 6 pg of sequences complementary to the probe in total RNA samples of 10 micrograms. A short-term time-course analysis showed that vitellogenin mRNA was detectable within 3 hr of exposure to estrogen during primary stimulation, and that the maximum rate of accumulation was reached at 5--6 hr. A long-term time-course analysis of the accumulation of vitellogenin mRNA showed that it is possible to obtain a primary response, a hormone withdrawal effect and an enhanced secondary response in the same batch of cells in a manner analogous to that observed in vivo. Measurement of hormone concentration dependence showed a response at 10(-9) M estradiol, which continued to increase up to at least 10(-6) M estradiol. This requirement for large doses of estradiol for maximal response can be explained by the rapid metabolism of estradiol by the cultured cells.