Tcf transcription factors interact with ␤-catenin and Armadillo to mediate Wnt/Wingless signaling. We now report the characterization of genes encoding two murine members of the Tcf family, mTcf-3 and mTcf-4. mTcf-3 mRNA is ubiquitously present in embryonic day 6.5 (E6.5) mouse embryos but gradually disappears over the next 3 to 4 days. mTcf-4 expression occurs first at E10.5 and is restricted to di-and mesencephalon and the intestinal epithelium during embryogenesis. The mTcf-3 and mTcf-4 proteins bind a canonical Tcf DNA motif and can complex with the transcriptional coactivator ␤-catenin. Overexpression of Wnt-1 in a mammary epithelial cell line leads to the formation of a nuclear complex between ␤-catenin and Tcf proteins and to Tcf reporter gene transcription. These data demonstrate a direct link between Wnt stimulation and ␤-catenin/Tcf transcriptional activation and imply a role for mTcf-3 and -4 in early Wnt-driven developmental decisions in the mouse embryo.

In many mammalian species, the intestinal epithelium undergoes major changes that allow a dietary transition from mother's milk to the adult diet at the end of the suckling period. These complex developmental changes are the result of a genetic programme intrinsic to the gut tube, but its regulators have not been identified. Here we show that transcriptional repressor B lymphocyte-induced maturation protein 1 (Blimp1) is highly expressed in the developing and postnatal intestinal epithelium until the suckling to weaning transition. Intestine-specific deletion of Blimp1 results in growth retardation and excessive neonatal mortality. Mutant mice lack all of the typical epithelial features of the suckling period and are born with features of an adult-like intestine. We conclude that the suckling to weaning transition is regulated by a single transcriptional repressor that delays epithelial maturation.

With respect to hepatocyte heterogeneity in ammonia and amino acid metabolism, two different patterns of sublobular gene expression are distinguished: ‘gradient-type’ and ‘strict- or compartment-type’ zonation. An example for strict-type zonation is the reciprocal distribution of carbamoylphosphate synthase and glutamine synthase in the liver lobule. The mechanisms underlying the different sublobular gene expressions are not yet settled but may involve the development of hepatic architecture, innervation, blood-borne hormonal and metabolic factors. The periportal zone is characterized by a high capacity for uptake and catabolism of amino acids (except glutamate and aspartate) as well as for urea synthesis and gluconeogenesis. On the other hand, glutamine synthesis, ornithine transamination and the uptake of vascular glutamate, aspartate, malate and a-ketoglutarate are restricted to a small perivenous hepatocyte population. Accordingly, in the intact liver lobule the major pathways for ammonia detoxication, urea and glutamine synthesis, are anatomically switched behind each other and represent in functional terms the sequence of the periportal low affinity system (urea synthesis) and a previous high affinity system (glutamine synthesis) for ammonia detoxication. Perivenous glutamine synthase-containing hepatocytes (‘scavenger cells’) act as a high affinity scavenger for the ammonia, which escapes the more upstream urea-synthesizing compartment. Periportal glutaminase acts as a pH- and hormone- modulated ammonia-amplifying system in the mitochondria of periportal hepatocytes. The activity of this amplifying system is one crucial determinant for flux through the urea cycle in view of the high K(m) (ammonia) of carbamoylphosphate synthase, the ratecontrolling enzyme of the urea cycle. The structural and functional organization of glutamine and ammonia-metabolizing pathways in the liver lobule provides one basis for the understanding of a hepatic role in systemic acid base homeostasis. Urea synthesis is a major pathway for irreversible removal of metabolically generated bicarbonate. The lobular organization enables the adjustment of the urea cycle flux and accordingly the rate of irreversible hepatic bicarbonate elimination to the needs of the systemic acid base situation, without the threat of hyperammonemia.

In order to maintain a stable karyotype, the eukaryotic cell cycle is coordinated such that only one round of S phase precedes each mitosis, and mitosis is not initiated until DNA replication is completed. Several checkpoints and regulatory proteins have been defined in lower eukaryotes that govern this coordination, but little is known about the proteins that are involved in mammalian cells. Previously, we have shown that the winged-helix transcription factor Trident - also known as HFH-11, FKL16 and WIN [1] [2] [3] - is exclusively expressed in cycling cells and is phosphorylated during mitosis [1] [4]. The cellular function of Trident has yet to be described, however. Here, we have shown that disruption of the Trident gene in mice resulted in postnatal death, most probably because of circulatory failure. Histological analysis of Trident -/- embryos from embryonic day 10 (E10) onwards revealed a specific, characteristic defect in the developing myocardium. The orientation of the myocytes was highly irregular and the nuclei of these disorganized cardiomyocytes were clearly polyploid with up to a 50-fold increase in DNA content. Polyploidy was also observed in embryonic hepatocytes. Our results indicate that expression of Trident is required to prevent multiple rounds of S phase in the heart and the liver. Trident therefore appears to have a role in preventing DNA re-replication during the G2 and M phases.

During development, the single-circuited cardiac tube transforms into a double-circuited four-chambered heart by a complex process of remodeling, differential growth, and septation. In this process the endocardial cushion tissues of the atrioventricular junction and outflow tract (OFT) play a crucial role as they contribute to the mesenchymal components of the developing septa and valves in the developing heart. After fusion, the endocardial ridges in the proximal portion of the OFT initially form a mesenchymal outlet septum. In the adult heart, however, this outlet septum is basically a muscular structure. Hence, the mesenchyme of the proximal outlet septum has to be replaced by cardiomyocytes. We have dubbed this process "myocardialization." Our immunohistochemical analysis of staged chicken hearts demonstrates that myocardialization takes place by ingrowth of existing myocardium into the mesenchymal outlet septum. Compared to other events in cardiac septation, it is a relatively late process, being initialized around stage H/H28 and being basically completed around stage H/H38. To unravel the molecular mechanisms that are responsible for the induction and regulation of myocardialization, an in vitro culture system in which myocardialization could be mimicked and manipulated was developed. Using this in vitro myocardialization assay it was observed that under the standard culture conditions (i) whole OFT explants from stage H/H20 and younger did not spontaneously myocardialize the collagen matrix, (ii) explants from stage H/H21 and older spontaneously formed extensive myocardial networks, (iii) the myocardium of the OFT could be induced to myocardialize and was therefore "myocardialization-competent" at all stages tested (H/H16-30), (iv) myocardialization was induced by factors produced by, most likely, the nonmyocardial component of the outflow tract, (v) at none of the embryonic stages analyzed was ventricular myocardium myocardialization-competent, and finally, (vi) ventricular myocardium did not produce factors capable of supporting myocardialization.