Rst1 and Rst2 repress the mating and filamentous growth responses of S. cerevisiae by directly inhibiting Ste12. Activation of Fus3 or Kss1 may cause phosphorylation-dependent release of Ste12 from Rst1/Rst2 and thereby activate Ste12-dependent transcription.

The liver-specific enhancer of the serum albumin gene contains an essential segment, designated eH, which binds the hepatocyte nuclear factor 3a (HNF3a) and ubiquitous nuclear factor 1/CCAAT transcription factor (NF1/CTF) proteins in tight apposition. We previously showed that activation of transcription by the eH site was correlated with an increase in intracellular HNF3ae levels during the in vitro differentiation of the hepatic cell line H2.35. We now show that transfection of an HNF3oa cDNA expression vector into dedifferentiated H2.35 cells is sufficient to induce transcription from the eH site. Mutational analysis of the enhancer demonstrates that NF1/CTF cooperates with HNF3a to induce enhancer activity. However, when the eH site is removed from the context of the enhancer, NF1/CTF can inhibit transcriptional activation by HNF3a. We conclude that the ternary complex of HNF3a, NF1/CTF, and the eH site forms a novel, composite regulatory element that is sensitive to the local DNA sequence environment and suggest that the transcriptional stimulatory activity of NF1/CTF depends on its higher-order interactions with other proteins during hepatocyte differentiation. Cellular differentiation is governed by the binding of transcription factors to DNA in specific combinations, often leading to the formation of large arrays of protein-DNA complexes. Interactions between different transcription factors bound to adjacent sites can lead to striking changes in their respective activities. For example, the yeast protein MCM1 (PRTF) activates a-cell-specific promoters when bound adjacent to the al transcription factor, but MCM1 represses a-cell-specific promoters when bound next to the a2 protein (1,29). In this report, we study the interactions between two liver transcription factors when they are tightly juxtaposed at a site on DNA and explore how the genetic context of the proteins' DNA binding sites governs their interactions during hepatic differentiation.Serum albumin gene transcription is an excellent marker for liver differentiation in mammals because it is activated early during liver development (8,49,51,53) and increases to a rate 1,000-fold greater in the liver than in other tissues (33,44). Tissue specificity is controlled in part by an enhancer element that lies 10 kb upstream of the transcription start site and functions selectively in the liver of transgenic mice (43). The albumin enhancer contains three sites, designated eE, eG, and eH, that are essential for enhancer activity in various hepatocyte-derived cell lines, such as H2.35 cells (24,25,34). The eE site binds liver-enriched C/EBP-related proteins (6, 11, 56), and the eG site binds a family of transcription factors, hepatocyte nuclear factor 3a (HNF3a), HNF3,, and HNF3y, expressed in liver and lung (31). The HNF3 family contains a conserved DNA binding domain found in the developmentally important Drosophila fork head protein (54). The eH site was defined by a large DNase I footprint with nuclear extracts from mouse liver and H2.35 cells (34,58); ...

Little is known about genes that govern the development of the definitive endoderm in mammals; this germ layer gives rise to the intestinal epithelium and various other cell types, such as hepatocytes, derived from the gut. The discovery that the rat hepatocyte transcription factor HNF3 is similar to the Drosophila forkhead gene, which plays a critical role in gut development in the fly, led us to isolate genes containing the HNF3/forkhead (HFH) domain that are expressed in mouse endoderm development. We recovered mouse HNF3 beta from an embryo cDNA library and found that the gene is first expressed in the anterior portion of the primitive streak at the onset of gastrulation, in a region where definitive endoderm first arises. Its expression persists in axial structures derived from the mouse equivalent of Hensen's node, namely definitive endoderm and notochord, and in the ventral region of the developing neural tube. Expression of the highly related gene, HNF3 alpha, appears to initiate later than HNF3 beta and is first seen in midline endoderm cells. Expression subsequently appears in notochord, ventral neural tube, and gut endoderm in patterns similar to HNF3 beta. Microscale DNA binding assays show that HNF3 proteins are detectable in the midgut at 9.5 days p.c. At later stages HNF3 mRNAs and protein are expressed strongly in endoderm-derived tissues such as the liver. HNF3 is also the only known hepatocyte-enriched transcription factor present in a highly de-differentiated liver cell line that retains the capacity to redifferentiate to the hepatic phenotype. Taken together, these studies suggest that HNF3 alpha and HNF3 beta are involved in both the initiation and maintenance of the endodermal lineage. We also discovered a novel HFH-containing gene, HFH-E5.1, that is expressed transiently in posterior ectoderm and mesoderm at the primitive streak stage, and later predominantly in the neural tube. HFH-E5.1 is highly similar in structure and expression profile to the Drosophila HFH gene FD4, suggesting that HFH family members have different, evolutionarily conserved roles in development.