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Kenneth S Zaret

Publications and source records attributed to Kenneth S Zaret.

At least 19 recordsLinked to original sources
PubMed2006

Specific interactions of the wing domains of FOXA1 transcription factor with DNA.

FOX (forkhead box) transcription factors have diverse regulatory roles in development, signaling, and longevity, as well as being able to bind stably to target sites in silent chromatin. Crystal structure analysis showed that the FOXA DNA binding domain folds into a helix-turn-helix (HTH) motif flanked on either side by "wings" of polypeptide chain. The wings have the potential to interact with the DNA minor groove along the long axis of the DNA helix, flanking the HTH interactions with the major groove. Diverse FOX family homologs exist, and structural studies with certain DNA target sites suggest that neither of the wing regions are well ordered or provide a stable contribution to DNA target site binding. However, FOXA1 binds certain DNA target sites with high affinity, and as a monomer. To determine whether the wing domains contribute to stable DNA binding, we assessed complexes of FOXA with high and lower affinity DNA target sites by hydroxyl radical footprinting and site-directed mutagenesis. The data revealed clear protections predicted for wing interactions at the high affinity target, but less so at the lower affinity target, indicating that the wing domains stably interact with high affinity DNA sites for FOXA proteins.

Base Sequence↗
PubMed2006

An endothelial-mesenchymal relay pathway regulates early phases of pancreas development.

Understanding the tissue interactions that induce pancreatic progenitor cells from the embryonic endoderm provides insights into congenital malformations, tissue repair, and differentiating stem cells to a pancreatic fate. The specification of pancreatic progenitors within the dorsal endodermal epithelium has been thought to involve two phases of mesodermal interactions; first with the lateral plate mesoderm and notochord and then with aortic endothelial cells. Afterwards, branching morphogenesis of the pancreatic bud is induced by Isl-1-positive dorsal mesenchyme cells, whose growth is stimulated by factors in the circulation. Using mouse genetic models and embryo tissue explants, we show that the aortic endothelium and dorsal mesenchyme each possess an additional role in pancreatic induction, prior to the branching morphogenesis step. Specifically, we find that aortic endothelial cells promote the survival of nearby, Isl-1-positive dorsal mesenchyme, independently of factors from the circulation. Furthermore, we find that FGF10 signaling from the mesenchyme cells maintains Ptf1a expression in the dorsal pancreatic bud and appears genetically redundant with a role for the transcription factor gene HNF6 in promoting the induction of Pdx-1-positive dorsal endoderm. Together, these studies reveal a relay pathway from aortic endothelium to dorsal mesenchyme and then to the endoderm, along with functions of the dorsal mesenchyme that promote the initial differentiation of the dorsal pancreatic endoderm, prior to organ morphogenesis.

Animals↗
PubMed2006

Pancreatic beta cells: responding to the matrix.

In a recent issue of Developmental Cell, Nikolova et al. (2006) make the unusual discovery that, unlike most epithelial cells, pancreatic beta cells do not produce a basement membrane of extracellular matrix. Rather, the matrix is provided to a subset of beta cells by local capillary endothelial cells, explaining in part how the endothelium helps promote optimal levels of insulin production.

Animals↗
PubMed2006

An FGF response pathway that mediates hepatic gene induction in embryonic endoderm cells.

While particular combinations of mesodermal signals are known to induce distinct tissue-specific programs in the endoderm, there is little information about the response pathways within endoderm cells that control their specification. We have used signaling inhibitors on embryo tissue explants and whole-embryo cultures as well as genetic approaches to reveal part of an intracellular network by which FGF signaling helps induce hepatic genes and stabilize nascent hepatic cells within the endodermal epithelium. Specifically, we found that hepatic gene induction is elicited by an FGF/MAPK pathway. Although the PI3K pathway is activated in foregut endoderm cells, its inhibition does not block hepatic gene induction in explants; however, it does block tissue growth. We also found that at the onset of hepatogenesis, the FGF/MAPK and PI3K pathways do not crossregulate in the endoderm. The finding of separate pathways for endoderm tissue specification and growth provides insights for guiding cellular regeneration and stem cell differentiation.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Hex homeobox gene controls the transition of the endoderm to a pseudostratified, cell emergent epithelium for liver bud development.

Little is known about the mechanism by which embryonic liver, lung, and pancreas progenitor cells emerge from the endodermal epithelium to initiate organogenesis. Understanding this process and its genetic control provides insight into ontogeny, developmental abnormalities, and tissue regeneration. We find that shortly after hepatic endoderm cells are specified, they undergo a transition from a columnar, gut morphology to a pseudostratified morphology, with concomitant "interkinetic nuclear migration" (INM) during cell division. INM is a hallmark of pseudostratified epithelia and the process used by neural progenitors to emerge from the neural epithelium. We find that the transition of the hepatic endoderm, but not the neural epithelium, to a pseudostratified epithelium is dependent upon the cell-autonomous activity of the homeobox gene Hex. In the absence of Hex, hepatic endoderm cells survive but maintain a columnar, simple epithelial phenotype and ectopically express Shh and other genes characteristic of the midgut epithelium. Thus, Hex promotes endoderm organogenesis by promoting the transition to a pseudostratified epithelium, which in turn allows hepatoblasts to emerge into the stromal environment and continue differentiating.

Animals↗
PubMed2005

Distinct populations of endoderm cells converge to generate the embryonic liver bud and ventral foregut tissues.

The location and movement of mammalian gut tissue progenitors, prior to the expression of tissue-specific genes, has been unknown, but this knowledge is essential to identify transitions that lead to cell type specification. To address this, we used vital dyes to label exposed anterior endoderm cells of early somite stage mouse embryos, cultured the embryos into the tissue bud phase of development, and determined the tissue fate of the dye labeled cells. This approach was performed at three embryonic stages that are prior to, or coincident with, foregut tissue patterning (1-3 somites, 4-6 somites, and 7-10 somites). Short-term labeling experiments tracked the movement of tissue progenitor cells during foregut closure. Surprisingly, we found that two distinct types of endoderm-progenitor cells, lateral and medial, arising from three spatially separated embryonic domains, converge to generate the epithelial cells of the liver bud. Whereas the lateral endoderm-progenitors give rise to descendants that are constrained in tissue fate and position along the anterior-posterior axis of the gut, the medial gut endoderm-progenitors give rise to descendants that stream along the anterior-posterior axis at the ventral midline and contribute to multiple gut tissues. The fate map reveals extensive morphogenetic movement of progenitors prior to tissue specification, it permits a detailed analysis of endoderm tissue patterning, and it illustrates that diverse progenitor domains can give rise to individual tissue cell types.

Animals↗
PubMed2004

FoxA1 binding to the MMTV LTR modulates chromatin structure and transcription.

Novel binding sites for the forkhead transcription factor family member Forkhead box A (FoxA), previously referred to as Hepatocyte Nuclear Factor 3 (HNF3), were found within the mouse mammary tumor virus long terminal repeat (MMTV LTR). The effect of FoxA1 on MMTV LTR chromatin structure, and expression was evaluated in Xenopus laevis oocytes. Mutagenesis of either of the two main FoxA binding sites showed that the distal site, -232/-221, conferred FoxA1-dependent partial inhibition of glucocorticoid receptor (GR) driven MMTV transcription. The proximal FoxA binding segment consisted of two individual FoxA sites at -57/-46 and -45/-34, respectively, that mediated an increased basal MMTV transcription. FoxA1 binding altered the chromatin structure of both the inactive- and the hormone-activated MMTV LTR. Hydroxyl radical foot printing revealed FoxA1-mediated changes in the nucleosome arrangement. Micrococcal nuclease digestion showed the hormone-dependent sub-nucleosome complex, containing approximately 120 bp of DNA, to be expanded by FoxA1 binding to the proximal segment into a larger complex containing approximately 200 bp. The potential function of the FoxA1-mediated expression of the MMTV provirus for maintenance of expression in different tissues is discussed.

Animals↗
PubMed2004

miR-122, a mammalian liver-specific microRNA, is processed from hcr mRNA and may downregulate the high affinity cationic amino acid transporter CAT-1.

These studies show that miR-122, a 22-nucleotide microRNA, is derived from a liver-specific noncoding polyadenylated RNA transcribed from the gene hcr. The exact sequence of miR-122 as well as the adjacent secondary structure within the hcr mRNA are conserved from mammalian species back to fish. Levels of miR-122 in the mouse liver increase to half maximal values around day 17 of embryogenesis, and reach near maximal levels of 50,000 copies per average cell before birth. Lewis et al. (2003) predicted the cationic amino acid transporter (CAT-1 or SLC7A1) as a miR-122 target. CAT-1 protein and its mRNA are expressed in all mammalian tissues but with lower levels in adult liver. Furthermore, during mouse liver development CAT-1 mRNA decreases in an almost inverse correlation with miR-122. Eight potential miR-122 target sites were predicted within the human CAT-1 mRNA, with six in the 3'-untranslated region. Using a reporter construct it was found that just three of the predicted sites, linked in a 400-nucleotide sequence from human CAT-1, acted with synergy and were sufficient to strongly inhibit protein synthesis and reduce mRNA levels. In summary, these studies followed the accumulation during development of miR-122 from its mRNA precursor, hcr, through to identification of what may be a specific mRNA target, CAT-1.

Animals↗
PubMed2004

Endothelial cell interactions initiate dorsal pancreas development by selectively inducing the transcription factor Ptf1a.

Dorsal and ventral pancreatic bud development from the endoderm requires inductive interactions with diverse mesodermal cell types and the action of transcription factors expressed within the endoderm. Presently it is unclear which mesodermal interactions activate which pancreatic transcription factors, and whether such inductions are common for initiating dorsal and ventral pancreas development. Previous studies of Lammert et al. showed that signaling from embryonic blood vessel cells, derived from the mesoderm, promotes pancreatic bud development. Using a combination of mouse Flk1(-/-) embryos lacking endothelial cells and tissue recombination experiments, we discovered that the initial induction of dorsal endoderm cells positive for the pancreatic and duodenal transcription factor Pdx1 does not require aorta or endothelial cell interactions, but dorsal pancreatic bud emergence and the maintenance of Pdx1 expression does. Aortal endothelial cells induce the crucial pancreatic transcription factor Ptf1a in the dorsal pancreatic endoderm; whereas the vitelline veins, which are normally adjacent to the emerging ventral pancreatic bud, are unnecessary for ventral Ptf1a induction or for ventral pancreatic bud initiation. We find that the aorta cells themselves, apart from the blood supply, cause the induction of Ptf1a in dorsal endoderm explants. Thus, endothelial cell interactions specifically promote early dorsal pancreatic development, at least in part, by inducing Ptf1a(+) pancreatic progenitors. Additionally, we find that endothelial cells are necessary for the induction of both the insulin and glucagon genes.

Animals↗
PubMed2004

Hex homeobox gene-dependent tissue positioning is required for organogenesis of the ventral pancreas.

In animal development, digestive tissues emerge from different positions of the endoderm as a result of patterning signals from overlying mesoderm. Although embryonic tissue movement during gastrulation generates an initial positional relationship between the endoderm and mesoderm, the role of subsequent endoderm movement against the mesoderm in patterning is unknown. At embryonic day 8.5 in the mouse, proliferation of cells at the leading edge of ventral-lateral endoderm, where the liver and ventral pancreas emerge, helps close off the foregut. During this time, the endoderm grows adjacent to and beyond the cardiogenic mesoderm, an inducer of the liver program and an inhibitor of the pancreas program. The homeobox gene Hex is expressed in this endoderm cell domain and in the liver and ventral pancreas buds, after organogenesis. We have found that in Hex(-/-) embryos, there is a complete failure in ventral pancreatic specification, while the liver program is still induced. However, when Hex-null ventral endoderm is isolated prior to its interaction with cardiogenic mesoderm and is cultured in vitro, it activates early pancreas genes. We found that Hex controls the proliferation rate, and thus the positioning, of the leading edge of endoderm cells that grow beyond the cardiogenic mesoderm, during gut tube closure. Thus, Hex-controlled positioning of endoderm cells beyond cardiogenic mesoderm dictates ventral pancreas specification. Other endodermal transcription factors may also function morphogenetically rather than by directly regulating tissue-specific programs.

Animals↗
PubMed2004

Liver development update: new embryo models, cell lineage control, and morphogenesis.

The three phases of liver development that are the focus of this review are: the specification of hepatoblasts within the endoderm, the lineage split of hepatoblasts into hepatocytes and biliary cells, and the interaction of these cells with different mesodermal cell derivatives during liver morphogenesis. Advances in these areas include new genes and experimental models for studying liver development, the role of HNF6 and HNF1beta transcription factors and notch signaling in the hepatocyte-biliary cell lineage decision, the identification of genomic targets for HNF4, and HNF4's role in controlling hepatic epithelial structure and the sinusoidal organization of the liver.

Animals↗
PubMed2004

Integrase-specific enhancement and suppression of retroviral DNA integration by compacted chromatin structure in vitro.

Integration of viral DNA into the host chromosome is an obligatory step in retroviral replication and is dependent on the activity of the viral enzyme integrase. To examine the influence of chromatin structure on retroviral DNA integration in vitro, we used a model target comprising a 13-nucleosome extended array that includes binding sites for specific transcription factors and can be compacted into a higher-ordered structure. We found that the efficiency of in vitro integration catalyzed by human immunodeficiency virus type 1 (HIV-1) integrase was decreased after compaction of this target with histone H1. In contrast, integration by avian sarcoma virus (ASV) integrase was more efficient after compaction by either histone H1 or a high salt concentration, suggesting that the compacted structure enhances this reaction. Furthermore, although site-specific binding of transcription factors HNF3 and GATA4 blocked ASV DNA integration in extended nucleosome arrays, local opening of H1-compacted chromatin by HNF3 had no detectable effect on integration, underscoring the preference of ASV for compacted chromatin. Our results indicate that chromatin structure affects integration site selection of the HIV-1 and ASV integrases in opposite ways. These distinct properties of integrases may also affect target site selection in vivo, resulting in an important bias against or in favor of integration into actively transcribed host DNA.

Avian Sarcoma Viruses↗
PubMed2003

Mouse Mix gene is activated early during differentiation of ES and F9 stem cells and induces endoderm in frog embryos.

In frog and zebrafish, the Mix/Bix family of paired type homeodomain proteins play key roles in specification and differentiation of mesendoderm. However, in mouse, only a single Mix gene (mMix) has been identified to date and its function is unknown. We have analyzed the expression of mouse Mix RNA and protein in embryos, embryoid bodies formed from embryonic stem cells and F9 teratocarcinoma cells, as well as several differentiated cell types. Expression in embryoid bodies in culture mirrors that in embryos, where Mix is transcribed transiently in primitive (visceral) endoderm (VE) and in nascent mesoderm. In F9 cells induced by retinoic acid to differentiate to VE, mMix is coordinately expressed with three other endodermal transcription factors, well before AFP, and its protein product is localized to the nucleus. In a subpopulation of nascent mesodermal cells from embryonic stem cell embryoid bodies, mMix is coexpressed with Brachyury. Intriguingly, mMix mRNA is detected in a population (T+Flk1+) of cells which may contain hemangioblasts, before the onset of hematopoiesis and activation of hematopoietic markers. In vitro and in vivo, mMix expression in nascent mesoderm is rapidly down-regulated and becomes undetectable in differentiated cell types. In the region of the developing gut, mMix expression is confined to the mesoderm of mid- and hindgut but is absent from definitive endoderm. Injection of mouse mMix RNA into early frog embryos results in axial truncation of developing tadpoles and, in animal cap assays, mMix alone is sufficient to activate expression of several endodermal (but not mesodermal) markers. Although these observations do not exclude a possible cell-autonomous function for mMix in mesendodermal progenitor cells, they do suggest an additional, non-cell autonomous role in nascent mesoderm in the formation and/or patterning of adjacent definitive endoderm.

Animals↗
PubMed2003

Hepatocyte nuclear factor 4alpha controls the development of a hepatic epithelium and liver morphogenesis.

Although advances have been made in understanding cell differentiation, only rudimentary knowledge exists concerning how differentiated cells form tissues and organs. We studied liver organogenesis because the cell and tissue architecture of this organ is well defined. Approximately 60% of the adult liver consists of hepatocytes that are arranged as single-cell anastomosing plates extending from the portal region of the liver lobule toward the central vein. The basal surface of the hepatocytes is separated from adjacent sinusoidal endothelial cells by the space of Disse, where the exchange of substances between serum and hepatocytes takes place. The hepatocyte's apical surface forms bile canaliculi that transport bile to the hepatic ducts. Proper liver architecture is crucial for hepatic function and is commonly disrupted in disease states, including cirrhosis and hepatitis. Here we report that hepatocyte nuclear factor 4alpha (Hnf4alpha) is essential for morphological and functional differentiation of hepatocytes, accumulation of hepatic glycogen stores and generation of a hepatic epithelium. We show that Hnf4alpha is a dominant regulator of the epithelial phenotype because its ectopic expression in fibroblasts induces a mesenchymal-to-epithelial transition. Most importantly, the morphogenetic parameters controlled by Hnf4alpha in hepatocytes are essential for normal liver architecture, including the organization of the sinusoidal endothelium.

Animals↗
PubMed2003

Insulation of the chicken beta-globin chromosomal domain from a chromatin-condensing protein, MENT.

Active genes are insulated from developmentally regulated chromatin condensation in terminally differentiated cells. We mapped the topography of a terminal stage-specific chromatin-condensing protein, MENT, across the active chicken beta-globin domain. We observed two sharp transitions of MENT concentration coinciding with the beta-globin boundary elements. The MENT distribution profile was opposite to that of acetylated core histones but correlated with that of histone H3 dimethylated at lysine 9 (H3me2K9). Ectopic MENT expression in NIH 3T3 cells caused a large-scale and specific remodeling of chromatin marked by H3me2K9. MENT colocalized with H3me2K9 both in chicken erythrocytes and NIH 3T3 cells. Mutational analysis of MENT and experiments with deacetylase inhibitors revealed the essential role of the reaction center loop domain and an inhibitory affect of histone hyperacetylation on the MENT-induced chromatin remodeling in vivo. In vitro, the elimination of the histone H3 N-terminal peptide containing lysine 9 by trypsin blocked chromatin self-association by MENT, while reconstitution with dimethylated but not acetylated N-terminal domain of histone H3 specifically restored chromatin self-association by MENT. We suggest that histone H3 modification at lysine 9 directly regulates chromatin condensation by recruiting MENT to chromatin in a fashion that is spatially constrained from active genes by gene boundary elements and histone hyperacetylation.

3T3 Cells↗