Hormone-dependent Accumulation of Rough Endoplasmic Reticulum in Mouse Mammary Epithelial Cells in Vitro

Mammary epithelial cell differentiation, as effected by the interplay of insulin, corticosteroid(s), and prolactin, has been further studied in vitro. Emphasis has been placed on the synthesis of casein in relation to the fabrication and function of the rough endoplasmic reticulum (RER). Minimal hormone requirements for the accumulation of the RER are insulin and corticosteroid. This is based on determinations of membrane-linked NADH-cytochrome c reductase (EC 1.6.99.3) and of RNA content of isolated RER. Accumulation of the enzyme is affected very little by insulin, modestly increased by hydrocortisone, and greatly enhanced by the combination of the two hormones. RNA content of RER is virtually unaltered by hydrocortisone, but is increased by insulin. This effect of insulin is, however, enhanced by the presence of hydrocortisone. Addition of hydrocortisone to an insulin-containing postmitotic system causes redistribution of free ribosomes to the membrane-bound variety without a net increase in ribosomes. In the absence of the steroid, insulin promotes development of an unstable RER which also is deficient in NADH-cytochrome c reductase. Prolactin does not affect the membrane-linked enzyme. It does, however, stimulate the rate of RNA synthesis and increases the RNA content of the membrane system. In the absence of RER, prolactin also increases the rate of RNA synthesis, but this effect is not sustained. Three lines of evidence strongly suggest that casein formation by these epithelial cells is dependent upon the presence of the RER that is induced by insulin and hydrocortisone. (a) There is close agreement between casein synthesis and RER formation as a function of hydrocortisone concentration. (b) RER formation precedes casein synthesis. (c) Casein formation requires sustained stimulation of RNA synthesis by prolactin, which, in turn, is dependent on the presence of RER.