Expression of differentiation-specific proteins in preneoplastic mammary tissues in BALB/c mice

Abstract

The levels of milk-specific mRNAs (alpha-casein, whey acidic protein) and proteins (casein, alpha-lactalbumin) were examined in the DIM series of mammary preneoplastic outgrowth lines and tumors. The constitutive production of casein protein was variable among the preneoplastic DIM outgrowth lines maintained in virgin mice. Outgrowth line DIM-2 consistently expressed a very low level of casein mRNA and protein but no detectable alpha-lactalbumin protein. Outgrowth line DIM-4 expressed a high level of casein protein and no alpha-lactalbumin; however, by transplant generation 9, the levels of casein mRNA and protein were significantly decreased but remained greater than those found in DIM-2. Outgrowth line DIM-3 expressed high levels of casein protein during all transplant generations examined and sporadically exhibited detectable amounts of alpha-lactalbumin. The level of beta-casein mRNA in DIM-3 was 7 times greater than seen in DIM-4 outgrowths, but still only 2% of that measured in the normal mammary gland from lactating mice. The majority of tumors derived from the DIM outgrowth lines expressed very low levels of beta-casein mRNA and total casein protein, although occasionally tumors were observed with very high levels of casein expression. Immunoblot analysis of cellular extracts indicated that alpha, beta-, and gamma-caseins were expressed in the three outgrowth lines to varying degrees. Whey acidic protein mRNA attained barely detectable levels in the best of cases. Only outgrowth line DIM-3 responded to a normal lactogenic stimulus (lactating mouse) with significantly increased levels of milk-specific products. As determined by avidin-biotin peroxidase staining, the percentage of alveoli expressing beta-casein and alpha-lactalbumin proteins in lactating mice increased from 20 and 0%, respectively, in virgin mice to 85 and 40%, respectively, in lactating mice. Similarly, the levels of mRNA for beta-casein and whey acidic protein increased 8- and 5.5-fold, respectively. These results indicate that the cell populations selected by serial transplantation of preneoplastic mammary tissues fall into at least three categories with respect to expression of mammary-specific differentiation products: uninducible, inducible, constitutive. The DIM-3 outgrowth line appears to represent a highly inducible cell population. As concluded in earlier investigations, there was no correlation between secretory activity, morphology of the outgrowths, and tumor-producing capabilities in virgin mice. The effect of a normal lactogenic stimulus on the tumor potential of the DIM-3 line is currently under investigation.