Our studies have demonstrated the differential induction of cytochrome P-450 monooxygenase enzymes in the tissues of the rat placenta by the polyaromatic compounds beta-naphthoflavone (BNF) and 3-methlcholanthrene (3MC). Further studies with explants of isolated placental tissues show that the profile of (3H)-leucine incorporation into secreted proteins is altered in basal zone and yolk sac tissues from BetaNF/3MC-treated animals. The proposed research will seek to identify the proteins affected by betaNF/3MC as well as characterize biochemical alterations in the intracellular processes involved in their respective synthesis and secretion. The first line of investigation will examine whether a new major 40,000 (40K) molecular weight (M/r) protein present in the culture media from yolk sac explants is unique to betaNF or, rather, is a marker for Ah inductin by betaNF/3MC. Effects of betaNF/3MC on the incorporation of radiolabeled amino acids into 40K protein and alpha-fetoprotein (AFP), the main secretory product of the yolk sac, will be examined in culture media and tissues using 2-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and immunoblot procedures. Yolk sac mRNA will be characterized in cell-free translocation system, as well as by Northern blot analysis using a rat AFP cDNA probe. If a common mechanism is implicated in the BetaNF/3MC-related presence of the 40K protein, further studies will determine whether the synthesis of the 40K protein is realted, quantitatively and temporally, to the induction of ethocyresorufin-O-deethylase (EROD) activity by BetaNF, 3MC and TCDD. The next aim will be to evaluate whether BetaNF/3MC-treatments have decreased AFP secretion from yolk sac through alterations in post-translational glycosylation mechanisms. AFP molcular heterogeneity will be characterized by Con A-Sepharose chromatography and 2D-PAGE. A separate line of investigation will study explants of basal zone tissue to determine if BetaNF/3MC treatments selectively alter the synthesis and secretion of rat placental lactogen (PL) or a complex of other 20-30K M/r proteins. The final goal is to establish whether concentrations of AFP and AP in maternal serum are altered by BetaNF/3MC and, moreover, correlate with placental and fetal weight. In summary, the proposed research will characterize biochemical alterations in the synthesis of secretory proteins in placental tissue from betaNF/3MC-treated rats. The results wil further help to elucidate whether induction of Ah is an indicator of toxicity in palcental function.
