A complex noncoordinate regulation of alpha-lactalbumin and 25 K beta-casein by corticosterone, prolactin, and insulin in long term cultures of normal rat mammary cells.

The concentrations of PRL, corticosterone, and insulin required by long term cultures of normal rat mammary cells to produce alpha-lactalbumin (alpha LA) and the 25,000 mol wt beta-casein were evaluated with a variety of hormone ratios and concentrations. For these studies a double antibody RIA for beta-casein capable of measuring 0.5 ng beta-casein/100 microliter growth media was developed and used along with our previously reported RIA for alpha LA. PRL was active at physiological levels (0.05-0.15 micrograms/ml) and quantitatively stimulated beta-casein more than alpha LA, whereas physiological levels of corticosterone (0.05-0.15 micrograms/ml) quantitatively stimulated alpha LA more than beta-casein. The concentration of corticosterone greatly altered the magnitude of the cells' response to insulin and PRL for alpha LA output by cells from either virginal or midpregnant rats. Insulin also enhanced production of these milk proteins, but very little effect was measured in the physiological range. alpha LA was increased more by insulin than by PRL, and beta-casein was enhanced more by insulin than by corticosterone. Cells from midpregnant rats required less insulin to stimulate beta-casein production than to stimulate alpha LA. Cells from virginal rats required a supraphysiological insulin level to stimulate both beta-casein and alpha LA under these conditions. These cells generally require 5-6 weeks to achieve a steady-state rate of milk protein output. The complexities of our observations help explain some of the conflicting reports in the literature concerning which hormone is of prime importance for quantitatively increasing the synthesis of a particular milk protein, particularly since high hormone levels are often employed and time in culture varies considerably among reports. We conclude that lower levels of all these hormones can and should be used in vitro. Our messenger RNA (mRNA) studies using cloned complementary DNA probes for two rat casein mRNAs show that cells grown for 2 months with hormones contain significant amounts of both alpha- and beta-casein mRNAs. Simultaneous quantification of beta-casein mRNA levels and rates of beta-casein protein production in these long term cell cultures indicated that a substantial portion of their beta-casein protein production is regulated by the amount of its mRNA. This could be controlled by mRNA synthesis and/or mRNA degradation.(ABSTRACT TRUNCATED AT 400 WORDS)