Freezing point depression, delta T/k, and pNa are measured and analyzed for aqueous solutions of trihydroxy (NaTC) and dihydroxy (NaDC and NaTDC) bile salts. The results show the existence of break points in the plot of delta T/k vs molality at 0.018, 0.013, and 0.007 m, respectively, in good agreement with previous published critical micelle concentration values. Above the break point bile salts form aggregates with average aggregation numbers of 2.59 +/- 0.12 (NaTC), 5.82 +/- 0.04 (NaDC), and 5.42 +/- 0.47 (NaTDC). Fractions of bound counterions are also deduced, being close to 0.3 for the three bile salts studied. This indicates that only one counterion is bound for every three monomers in the aggregate. The different structural models published for the bile salt aggregates are discussed.

Using chimeric recombinants transfected into HeLa cells and a transient expression assay, we demonstrate that the 5'-flanking region of the pS2 gene from position -428 to position -324 exhibits both constitutive and estrogeninducible enhancer activity. The estrogen-inducible activity, but not the constitutive activity, was inhibited by antiestrogens.ICI 164,384 behaved as a pure antagonist, whereas hydroxytamoxifen was a partial agonist-antagonist. The estrogenresponsive element of the pS2 gene has been narrowed down by site-directed deletion mutagenesis to a 13-base-pair (position -405 to position -393) imperfectly palindromic sequence, which in isolation can confer estrogen inducibility to the heterologous rabbit IB-globin gene promoter. On the other hand, the sequences responsible for the constitutive enhancer activity are spread over the entire region.The human pS2 gene was initially characterized as a gene whose expression is specifically controlled by estrogen in the breast cancer cell line MCF-7 (1, 2). The increase in pS2 mRNA after addition of estradiol to the culture medium is a primary transcriptional event (3), suggesting that control of pS2 gene promoter activity by the estrogen receptor is mediated by a cis-acting estrogen-responsive element (ERE) that could be located in the 5'-flanking region ofthe pS2 gene. Using MCF-7 cells transformed with chimeric recombinants containing the promoter region of the pS2 gene controlling transcription of the gene conferring resistance to neomycin (G418), it has indeed been shown that the 5'-flanking region of the pS2 gene from position approximately -3000 to position +10 contains an estrogen-responsive promoter (4). Transfection of these recombinants together with the human estrogen receptor expression vector HEO in the presence or absence of estradiol led to the hypothesis that both constitutive and estrogen-inducible enhancer elements are located within the region of the pS2 gene from position -428 to position -332 (7).In the present study we demonstrate that the pS2 gene sequence from position -428 to position -324 indeed contains both constitutive and estrogen-inducible enhancer elements. Furthermore, the exact location of the ERE has been determined by using both deletion and point mutagenesis. We show that the pS2 ERE sequence is related to those of characterized EREs but that it is not a perfect palindrome, which results in a weaker ERE. Fig. 3A (note the presence Abbreviation: ERE, estrogen-responsive element. MATERIALS AND METHODS 1218The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Expression of the pS2 gene which is transcriptionally controlled by oestrogens in the breast cancer cell line MCF‐7 is oestrogen independent in stomach mucosa. We show here that the level of MCF‐7 cell pS2 mRNA can also be increased by the tumour promoter 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA). We further demonstrate, using transient transfection assays, that the −428 to −332 5′ flanking sequence of the pS2 gene contains DNA enhancer elements responsive to oestrogens, TPA, EGF, the c‐Ha‐ras oncoprotein and the c‐jun protein.