The Evolutionary Origin of the Runx/CBFbeta Transcription Factors – Studies of the Most Basal Metazoans

BACKGROUND. Members of the Runx family of transcriptional regulators, which bind DNA as heterodimers with CBFβ, are known to play critical roles in embryonic development in many triploblastic animals such as mammals and insects. They are known to regulate basic developmental processes such as cell fate determination and cellular potency in multiple stem-cell types, including the sensory nerve cell progenitors of ganglia in mammals. RESULTS. In this study, we detect and characterize the hitherto unexplored Runx/CBFβ genes of cnidarians and sponges, two basal animal lineages that are well known for their extensive regenerative capacity. Comparative structural modeling indicates that the Runx-CBFβ-DNA complex from most cnidarians and sponges is highly similar to that found in humans, with changes in the residues involved in Runx-CBFβ dimerization in either of the proteins mirrored by compensatory changes in the binding partner. In situ hybridization studies reveal that Nematostella Runx and CBFβ are expressed predominantly in small isolated foci at the base of the ectoderm of the tentacles in adult animals, possibly representing neurons or their progenitors. CONCLUSION. These results reveal that Runx and CBFβ likely functioned together to regulate transcription in the common ancestor of all metazoans, and the structure of the Runx-CBFβ-DNA complex has remained extremely conserved since the human-sponge divergence. The expression data suggest a hypothesis that these genes may have played a role in nerve cell differentiation or maintenance in the common ancestor of cnidarians and bilaterians.National Science Foundation (IBN-0212773, FP-91656101-0); Boston University SPRInG (20-202-8103-9); Israel Science Foundation (825/07

Developmentally regulated promoter-switch transcriptionally controls Runx1 function during embryonic hematopoiesis

<p>Abstract</p> <p>Background</p> <p>Alternative promoters usage is an important paradigm in transcriptional control of mammalian gene expression. However, despite the growing interest in alternative promoters and their role in genome diversification, very little is known about how and on what occasions those promoters are differentially regulated. Runx1 transcription factor is a key regulator of early hematopoiesis and a frequent target of chromosomal translocations in acute leukemias. Mice deficient in <it>Runx1 </it>lack definitive hematopoiesis and die in mid-gestation. Expression of <it>Runx1 </it>is regulated by two functionally distinct promoters designated P1 and P2. Differential usage of these two promoters creates diversity in distribution and protein-coding potential of the mRNA transcripts. While the alternative usage of P1 and P2 likely plays an important role in <it>Runx1 </it>biology, very little is known about the function of the P1/P2 switch in mediating tissue and stage specific expression of <it>Runx1 </it>during development.</p> <p>Results</p> <p>We employed mice bearing a hypomorphic <it>Runx1 </it>allele, with a largely diminished P2 activity, to investigate the biological role of alternative P1/P2 usage. Mice homozygous for the hypomorphic allele developed to term, but died within a few days after birth. During embryogenesis the P1/P2 activity is spatially and temporally modulated. P2 activity is required in early hematopoiesis and when attenuated, development of liver hematopoietic progenitor cells (HPC) was impaired. Early thymus development and thymopoiesis were also abrogated as reflected by thymic hypocellularity and loss of corticomedullary demarcation. Differentiation of CD4/CD8 thymocytes was impaired and their apoptosis was enhanced due to altered expression of T-cell receptors.</p> <p>Conclusion</p> <p>The data delineate the activity of P1 and P2 in embryogenesis and describe previously unknown functions of Runx1. The findings show unequivocally that the role of P1/P2 during development is non redundant and underscore the significance of alternative promoter usage in Runx1 biology.</p

Cell type–specific actions of Bcl11b in early T-lineage and group 2 innate lymphoid cells

The zinc finger transcription factor, Bcl11b, is expressed in T cells and group 2 innate lymphoid cells (ILC2s) among hematopoietic cells. In early T-lineage cells, Bcl11b directly binds and represses the gene encoding the E protein antagonist, Id2, preventing pro-T cells from adopting innate-like fates. In contrast, ILC2s co-express both Bcl11b and Id2. To address this contradiction, we have directly compared Bcl11b action mechanisms in pro-T cells and ILC2s. We found that Bcl11b binding to regions across the genome shows distinct cell type–specific motif preferences. Bcl11b occupies functionally different sites in lineage-specific patterns and controls totally different sets of target genes in these cell types. In addition, Bcl11b bears cell type–specific post-translational modifications and organizes different cell type–specific protein complexes. However, both cell types use the same distal enhancer region to control timing of Bcl11b activation. Therefore, although pro-T cells and ILC2s both need Bcl11b for optimal development and function, Bcl11b works substantially differently in these two cell types

Absence of Runx3 expression in normal gastrointestinal epithelium calls into question its tumour suppressor function

The Runx3 transcription factor regulates cell fate decisions during embryonic development and in adults. It was previously reported that Runx3 is strongly expressed in embryonic and adult gastrointestinal tract (GIT) epithelium (Ep) and that its loss causes gastric cancer. More than 280 publications have based their research on these findings and concluded that Runx3 is indeed a tumour suppressor (TS). In stark contrast, using various measures, we found that Runx3 expression is undetectable in GIT Ep. Employing a variety of biochemical and genetic techniques, including analysis of Runx3-GFP and R26LacZ/Runx3Cre or R26tdTomato/Runx3Cre reporter strains, we readily detected Runx3 in GIT-embedded leukocytes, dorsal root ganglia, skeletal elements and hair follicles. However, none of these approaches revealed detectable Runx3 levels in GIT Ep. Moreover, our analysis of the original Runx3LacZ/LacZ mice used in the previously reported study failed to reproduce the GIT expression of Runx3. The lack of evidence for Runx3 expression in normal GIT Ep creates a serious challenge to the published data and undermines the notion that Runx3 is a TS involved in cancer pathogenesis

Cell type–specific actions of Bcl11b in early T-lineage and group 2 innate lymphoid cells

The zinc finger transcription factor, Bcl11b, is expressed in T cells and group 2 innate lymphoid cells (ILC2s) among hematopoietic cells. In early T-lineage cells, Bcl11b directly binds and represses the gene encoding the E protein antagonist, Id2, preventing pro-T cells from adopting innate-like fates. In contrast, ILC2s co-express both Bcl11b and Id2. To address this contradiction, we have directly compared Bcl11b action mechanisms in pro-T cells and ILC2s. We found that Bcl11b binding to regions across the genome shows distinct cell type–specific motif preferences. Bcl11b occupies functionally different sites in lineage-specific patterns and controls totally different sets of target genes in these cell types. In addition, Bcl11b bears cell type–specific post-translational modifications and organizes different cell type–specific protein complexes. However, both cell types use the same distal enhancer region to control timing of Bcl11b activation. Therefore, although pro-T cells and ILC2s both need Bcl11b for optimal development and function, Bcl11b works substantially differently in these two cell types

Runx3 and T-box proteins cooperate to establish the transcriptional program of effector CTLs

Activation of naive CD8+ T cells with antigen induces their differentiation into effector cytolytic T lymphocytes (CTLs). CTLs lyse infected or aberrant target cells by exocytosis of lytic granules containing the pore-forming protein perforin and a family of proteases termed granzymes. We show that effector CTL differentiation occurs in two sequential phases in vitro, characterized by early induction of T-bet and late induction of Eomesodermin (Eomes), T-box transcription factors that regulate the early and late phases of interferon (IFN) γ expression, respectively. In addition, we demonstrate a critical role for the transcription factor Runx3 in CTL differentiation. Runx3 regulates Eomes expression as well as expression of three cardinal markers of the effector CTL program: IFN-γ, perforin, and granzyme B. Our data point to the existence of an elaborate transcriptional network in which Runx3 initially induces and then cooperates with T-box transcription factors to regulate gene transcription in differentiating CTLs

Structure and regulated expression of mammalian RUNX genes. Oncogene 23:4211–4219

The RUNX are key regulators of lineage-specific gene expression in major developmental pathways. The expression of RUNX genes is tightly regulated, leading to a highly specific spatio/temporal expression pattern and to distinct phenotypes of gene knockouts. This review highlights the extensive structural similarities between the three mammalian RUNX genes and delineates how regulation of their expression at the levels of transcription and translation are orchestrated into the unique RUNX expression pattern

Runx3 knockouts and stomach cancer: The challenge of identifying phenotypic defects directly attributable to loss of gene function

Gene targeting often results in knockout mice that show several phenotypes, some of which may not directly relate to the intrinsic function of the disrupted gene. Hence, to study the biological function of genes using knockout mice, one must identify the defects that are directly due to the loss of the targeted gene. Runx3 is a transcription factor that regulates lineage-specific gene expression in developmental processes. Recently, two groups produced Runx3 knockout mice. Two comparable defects were identified in both knockout strains, one involved neurogenesis and the other thymopoiesis. In addition, a stomach defect pertaining to gastric cancer was observed in one of the mutant strains, but not in the other. Here, we assess the differences between the two Runx3 mutant strains and discuss further studies that could reconcile these discrepancies. This article highlights the difficulties of inferring gene function through the interpretation of knockout phenotypes

The False Paradigm of RUNX3 Function as Tumor Suppressor in Gastric Cancer

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