Theria-specific homeodomain and cis-regulatory element evolution of the Dlx3-4 bigene cluster in 12 different mammalian species.

The mammalian Dlx3 and Dlx4 genes are configured as a bigene cluster, and their respective expression patterns are controlled temporally and spatially by cis-elements that largely reside within the intergenic region of the cluster. Previous work revealed that there are conspicuously conserved elements within the intergenic region of the Dlx3-4 bigene clusters of mouse and human. In this paper we have extended these analyses to include 12 additional mammalian taxa (including a marsupial and a monotreme) in order to better define the nature and molecular evolutionary trends of the coding and non-coding functional elements among morphologically divergent mammals.

A yeast-based recombinogenic targeting toolset for transgenic analysis of human disease genes.

Transgenic mouse models are valuable resources for analyzing functions of genes involved in human diseases. Mouse models provide critical insights into biological processes, including in vivo visualization of vasculature critical to our understanding of the immune system. Generating transgenic mice requires the capture and modification of large-insert DNAs representing genes of interest.

Essential functions of the Williams-Beuren syndrome-associated TFII-I genes in embryonic development.

GTF2I and GTF2IRD1 encoding the multifunctional transcription factors TFII-I and BEN are clustered at the 7q11.23 region hemizygously deleted in Williams-Beuren syndrome (WBS), a complex multisystemic neurodevelopmental disorder. Although the biochemical properties of TFII-I family transcription factors have been studied in depth, little is known about the specialized contributions of these factors in pathways required for proper embryonic development.

Identification of the TFII-I family target genes in the vertebrate genome.

GTF2I and GTF2IRD1 encode members of the TFII-I transcription factor family and are prime candidates in the Williams syndrome, a complex neurodevelopmental disorder. Our previous expression microarray studies implicated TFII-I proteins in the regulation of a number of genes critical in various aspects of cell physiology. Here, we combined bioinformatics and microarray results to identify TFII-I downstream targets in the vertebrate genome.

Transgenic LacZ under control of Hec-6st regulatory sequences recapitulates endogenous gene expression on high endothelial venules.

Hec-6st is a highly specific high endothelial venule (HEV) gene that is crucial for regulating lymphocyte homing to lymph nodes (LN). The enzyme is also expressed in HEV-like vessels in tertiary lymphoid organs that form in chronic inflammation in autoimmunity, graft rejection, and microbial infection. Understanding the molecular nature of Hec-6st regulation is crucial for elucidating its function in development and disease.

Mouse naked cuticle 2 (mNkd2) as a direct transcriptional target of Hoxc8 in vivo.

Mouse naked cuticle 2 (mNkd2), the mammalian homolog of the Drosophila segment polarity gene naked cuticle (nkd), encodes an EF hand protein that regulates early Wg activity by acting as an inducible antagonist. The transcription factor, Hoxc8, a member of the homeobox gene family, is vital for growth and differentiation. Chromatin immunoprecipitation (ChIP) assay, an electrophoretic mobility shift assay (EMSA), and a reporter assay demonstrated that endogenous Hoxc8 protein binds directly to the enhancer region of the mNkd2 gene, implying a Hoxc8-dependent transcriptional activity.

Expression profiling of BEN regulated genes in mouse embryonic fibroblasts.

BEN is a member of the TFII-I family of helix-loop-helix transcription factors. Both TFII-I and BEN are involved in gene regulation through interactions with tissue-specific transcription factors and chromatin remodeling complexes. Identification of the downstream target genes of TFII-I proteins is critical in delineating the regulatory effects of these proteins.

Laser-assisted microdissection (LAM) in developmental biology.

The analysis of gene expression in developing organs is a valuable tool for the assessment of genetic fingerprints during the various stages of differentiation. Complex processes in developing tissues are particularly difficult to understand in terms of biochemical phenomena. Laser-assisted microdissection (LAM) allows the efficient and precise capture of cells or groups of cells from developing tissues in sufficient quantities and within the context of time and space to permit the subsequent molecular characterization of the targeted tissue.

Multiple roles of hoxc8 in skeletal development.

We are interested in investigating the function of Hoxc8 in skeletogenesis during mouse development. Previous studies have shown that deregulation of Hoxc8 expression in the mouse leads to several skeletal defects, such as homeotic transformation in the thoracic vertebrae, abnormal development of the rib cage, and overproliferation of chondrocytes in the hypertrophic area. By deleting a crucial enhancer of Hoxc8 in vivo, we found that precise temporal expression of Hoxc8 is important for determining the correct identity of the vertebral column in early embryos.

Identification of a Hoxc8-regulated transcriptional network in mouse embryo fibroblast cells.

The transcription factor, Hoxc8, is a member of the homeobox gene family that is vital for growth and differentiation. Previously, we identified 34 genes whose expression levels were changed at least 2-fold by forced expression of Hoxc8 in C57BL/6J mouse embryo fibroblast cells using a mouse 16,463-gene oligonucleotide microarray. In the present study, we used the combined power of microarray profiling, global Hoxc8 DNA-binding site analysis, and high-throughput chromatin immunoprecipitation assays to identify direct and biologically relevant targets of Hoxc8 in vivo.

HnRNP A3 genes and pseudogenes in the vertebrate genomes.

The hnRNP A/B type proteins are abundant nuclear factors that bind to Pol II transcripts and are involved in numerous RNA-related activities. To date most data on the hnRNP A/B family have been obtained with recombinant proteins and cell cultures. Further characterization can result from an examination of the impact of various modifications in intact functional loci; however, such characterization is hampered by the presence of numerous and widely dispersed hnRNP A/B-related sequences in the mammalian genome.

The identification of Hoxc8 target genes.

Hox genes encode transcription factors that control spatial patterning during embryogenesis. To date, downstream targets of Hox genes have proven difficult to identify. Here, we describe studies designed to identify target genes under the control of the murine transcription factor Hoxc8.

Evidence for independent Hox gene duplications in the hagfish lineage: a PCR-based gene inventory of Eptatretus stoutii.

Hox genes code for transcription factors that play a major role in the development of all animal phyla. In invertebrates these genes usually occur as tightly linked cluster, with a few exceptions where the clusters have been dissolved. Only in vertebrates multiple clusters have been demonstrated which arose by duplication from a single ancestral cluster.

GTF2IRD2 is located in the Williams-Beuren syndrome critical region 7q11.23 and encodes a protein with two TFII-I-like helix-loop-helix repeats.

Williams-Beuren syndrome (also known as Williams syndrome) is caused by a deletion of a 1.55- to 1.84-megabase region from chromosome band 7q11.

The shark HoxN cluster is homologous to the human HoxD cluster.

The statistical analysis of phylogenetic footprints in the two known horn shark Hox clusters and the four mammalian clusters shows that the shark HoxN cluster is HoxD-like. This finding implies that the most recent common ancestor of jawed vertebrates had at least four Hox clusters, including those which are orthologous to the four mammalian Hox clusters.

Bichir HoxA cluster sequence reveals surprising trends in ray-finned fish genomic evolution.

The study of Hox clusters and genes provides insights into the evolution of genomic regulation of development. Derived ray-finned fishes (Actinopterygii, Teleostei) such as zebrafish and pufferfish possess duplicated Hox clusters that have undergone considerable sequence evolution. Whether these changes are associated with the duplication(s) that produced extra Hox clusters is unresolved because comparison with basal lineages is unavailable.

The early embryonic expression of TFII-I during mouse preimplantation development.

We studied the developmentally regulated expression of mouse TFII-I, a founding member of a family of transcription factors characterized by the presence of multiple helix-loop-helix repeat domains. TFII-I and BEN, a second member of this family, are involved in histone modification and SUMOylation. The genes, GTF2I and GTF2IRD1, encoding these proteins in human are located at chromosomal band 7q11.

Homeoprotein DLX-1 interacts with Smad4 and blocks a signaling pathway from activin A in hematopoietic cells.

In the transforming growth factor beta (TGF-beta) superfamily, activin A, TGF-beta1, and bone morphogenic protein 4 (BMP-4) have various effects on hematopoiesis, including early mesodermo-hematogenesis. After these cytokines bind to their respective receptor, a regulatory Smad is phosphorylated and becomes associated with Smad4, the common Smad, and the resulting complex translocates to the nucleus to regulate transcription. DLX1 is the product of a member of the distal-less homeobox gene family, which is known to have important roles in embryogenesis, particularly in craniofacial development, and in GABAergic neurogenesis.

Hox cluster duplications and the opportunity for evolutionary novelties.

Hox genes play a key role in animal body plan development. These genes tend to occur in tightly linked clusters in the genome. Vertebrates and invertebrates differ in their Hox cluster number, with vertebrates having multiple clusters and invertebrates usually having only one.

Expression of BEN, a member of TFII-I family of transcription factors, during mouse pre- and postimplantation development.

BEN is a member of the TFII-I family of transcription factors, characterized by the presence of multiple helix-loop-helix repeat domains. Our immunohistochemical analysis demonstrated broad and extensive expression of BEN during mouse pre- and postimplantation development, with highest levels occurring during early to midgestation. Maternally expressed BEN is present in both the cytoplasm and nuclei of the zygote; however, it retains a predominantly nuclear localization between the two-cell stage of development and early blastocyst stages.

Homez, a homeobox leucine zipper gene specific to the vertebrate lineage.

This work describes a vertebrate homeobox gene, designated Homez (homeodomain leucine zipper-encoding gene), that encodes a protein with an unusual structural organization. There are several regions within Homez, including three atypical homeodomains, two leucine zipper-like motifs, and an acidic domain. The gene is ubiquitously expressed in human and murine tissues, although the expression pattern is more restricted during mouse development.

Enhancer timing of Hox gene expression: deletion of the endogenous Hoxc8 early enhancer.

The proper expression of Hox genes is necessary for the accurate patterning of the body plan. The elucidation of the developmental genetic basis of transcriptional regulation of Hox genes by the study of their cis-regulatory elements provides crucial information regarding the establishment of axial specification. In this report, we investigate the role of the early enhancer (EE) of the murine Hoxc8 gene to better understand its role in pattern formation.