No abstract

The winged helix transcription factor hepatocyte nuclear factor 3␥ (HNF3␥) is expressed in embryonic endoderm and its derivatives liver, pancreas, stomach, and intestine, as well as in testis and ovary. We have generated mice carrying an Hnf3g-lacZ fusion which deletes most of the HNF3␥ coding sequence as well as 5.5 kb of 3 flanking region. Mice homozygous for the mutation are fertile, develop normally, and show no morphological defects. The mild phenotype change of the Hnf3g ؊/؊ mice can be explained in part by an upregulation of HNF3␣ and HNF3␤ in the liver of the mutant animals. Analysis of steady-state mRNA levels as well as transcription rates showed that levels of expression of several HNF3 target genes (phosphoenolpyruvate carboxykinase, transferrin, tyrosine aminotransferase) were reduced by 50 to 70%, indicating that HNF3␥ is an important activator of these genes in vivo.Transcription factors control gene expression in adult liver and hepatoma cell lines. These factors contain various structural motifs to allow for high-specificity DNA binding; among them are the divergent homeodomain proteins hepatocyte nuclear factor 1␣ (HNF1␣) and HNF1␤; the winged helix proteins HNF3␣, -␤, and -␥; the leucine zipper proteins C/EBP␣ and -␤; the orphan nuclear receptor HNF4; and the D site binding protein (DBP) (reviewed in references 5 and 32). Liver-specific gene expression cannot be achieved through the action of any individual transcription factor, because none of these factors is expressed exclusively in liver. When cis-regulatory elements of liver-specific genes were analyzed in detail, binding sites for multiple transcription factors were found. Therefore, the combinatorial action of the regulatory proteins most likely produces the stringency of hepatic gene expression.The HNF3 proteins were discovered by their ability to bind to the promoters of the genes encoding ␣1-antitrypsin (␣1-AT) and transthyretin (TTR) (8). Cloning of the cDNAs identified three HNF3 genes (HNF3␣, -␤, and -␥) in mammals (14, 15). The HNF3 genes are closely related to the Drosophila melanogaster gene forkhead, which is essential for the proper formation of the foregut and hindgut in flies (30). Therefore, it has been suggested that the HNF3 genes function in mammalian liver and gut development (15). This hypothesis is supported by the observation that the HNF3 genes are expressed very early during the formation of definite endoderm, from which liver and gut are derived (3,19,27).During formation of the definite endoderm, HNF3␤ is activated first, followed by HNF3␣, and finally HNF3␥ (3,19,27). The three HNF3 genes have different anterior boundaries of expression in the definite endoderm, suggesting that they are involved in the regionalization of the primitive gut tube. In addition, HNF3␤ is expressed in the node, and HNF3␣ and HNF3␤ are both expressed in the notochord and floorplate (3,19,27). HNF3␤ is absolutely required for notochord and floorplate formation, because these structures are missing in embryos homozygous for a targeted n...

The enormous cellular complexity of the brain is a major obstacle for gene expression profiling of neurological disease models, because physiologically relevant changes of transcription in a specific neuronal subset are likely to be lost in the presence of other neurons and glia. We solved this problem in transgenic mice by labeling genetically defined cells with a nuclear variant of GFP. When combined with laser-directed microdissection, intact RNA from unfixed, freeze-dried sections can be isolated, which is a prerequisite for high-quality global transcriptome analysis. Here, we compared gene expression profiles between pyramidal motor neurons and pyramidal somatosensory neurons captured from layer V of the adult neocortex. One striking feature of motor neurons is the elevated expression of ribosomal genes and genes involved in ATP synthesis. This suggests a molecular adaptation of the upper motor neurons to longer axonal projections and higher electrical activity. These molecular signatures were not detected when cortical layers and microareas were analyzed in toto.Additionally, we used microarrays to determine the global mRNA expression profiles of microdissected Purkinje cells and cellularly complex cerebellar cortex microregions. In summary, our analysis shows that cellularly complex targets lead to averaged gene expression profiles that lack substantial amounts of cell type-specific information. Thus, cell type-restricted sampling strategies are mandatory in the CNS. The combined use of a genetic label with laser-microdissection offers an unbiased approach to map patterns of gene expression onto practically any cell type of the brain.

No abstract