Hormone binding and localization of the c-erb-A protein suggest that it is a receptor for thyroid hormone, a nuclear protein that binds to DNA and activates transcription. In contrast, the product of the viral oncogene v-erb-A is defective in binding the hormone but is still located in the nucleus.

Gain of Wnt signaling through beta-catenin has been ascribed a critical function in the stimulation of hematopoietic stem cell self-renewal, whereas loss of beta-catenin is reportedly dispensable for hematopoiesis. Here we have used conditional mouse genetics and transplantation assays to demonstrate that constitutive activation of beta-catenin blocked multilineage differentiation, leading to the death of mice. Blood cell depletion was accompanied by failure of hematopoietic stem cells to repopulate irradiated hosts and to differentiate into mature cells. Activation of beta-catenin enforced cell cycle entry of hematopoietic stem cells, thus leading to exhaustion of the long-term stem cell pool. Our data suggest that fine-tuned Wnt stimulation is essential for hematopoiesis and is thus critical for therapeutic hematopoietic stem cell population expansion.

DNA methylation is a dynamic epigenetic mark that undergoes extensive changes during differentiation of self-renewing stem cells. However, whether these changes are the cause or consequence of stem cell fate remains unknown. Here, we show that alternative functional programs of hematopoietic stem cells (HSCs) are governed by gradual differences in methylation levels. Constitutive methylation is essential for HSC self-renewal but dispensable for homing, cell cycle control and suppression of apoptosis. Notably, HSCs from mice with reduced DNA methyltransferase 1 activity cannot suppress key myeloerythroid regulators and thus can differentiate into myeloerythroid, but not lymphoid, progeny. A similar methylation dosage effect controls stem cell function in leukemia. These data identify DNA methylation as an essential epigenetic mechanism to protect stem cells from premature activation of predominant differentiation programs and suggest that methylation dynamics determine stem cell functions in tissue homeostasis and cancer.

Monocytes are circulating, short-lived mononuclear phagocytes, which in mice and man comprise two main subpopulations. Murine Ly6C monocytes display developmental plasticity and are recruited to complement tissue-resident macrophages and dendritic cells on demand. Murine vascular Ly6C monocytes patrol the endothelium, act as scavengers, and support vessel wall repair. Here we characterized population and single cell transcriptomes, as well as enhancer and promoter landscapes of the murine monocyte compartment. Single cell RNA-seq and transplantation experiments confirmed homeostatic default differentiation of Ly6C into Ly6C monocytes. The main two subsets were homogeneous, but linked by a more heterogeneous differentiation intermediate. We show that monocyte differentiation occurred through de novo enhancer establishment and activation of pre-established (poised) enhancers. Generation of Ly6C monocytes involved induction of the transcription factor C/EBPβ and C/EBPβ-deficient mice lacked Ly6C monocytes. Mechanistically, C/EBPβ bound the Nr4a1 promoter and controlled expression of this established monocyte survival factor.

The activation of many genes requires the concerted effort of two or more transcription factors. Although C/EBP beta is known to cooperate with Myb to induce transcription of the granulocyte-specific mim-1 gene, the molecular mechanism of this cooperativity is undefined. We show that the N terminus of the full-length C/EBP beta isoform, which is essential for induction of the mim-1 gene in chromatin, interacts specifically with the SWI/SNF complex. Grafting this domain onto Myb generates a chimeric activator that recruits SWI/SNF and induces mim-1 transcription in the absence of C/EBP beta. Interaction between C/EBP beta and SWI/SNF is essential for activating a subgroup of resident target genes in chromatin and may represent a major determinant of combinatorial gene regulation in eukaryotes.

The LAZ3͞BCL6 (lymphoma-associated zinc finger 3͞B cell lymphomas 6) gene frequently is altered in non-Hodgkin lymphomas. It encodes a sequence-specific DNA binding transcriptional repressor that contains a conserved N-terminal domain, termed BTB͞POZ (bric-à-brac tramtrack broad complex͞pox viruses and zinc fingers). Using a yeast two-hybrid screen, we show here that the LAZ3͞BCL6 BTB͞ POZ domain interacts with the SMRT (silencing mediator of retinoid and thyroid receptor) protein. SMRT originally was identified as a corepressor of unliganded retinoic acid and thyroid receptors and forms a repressive complex with a mammalian homolog of the yeast transcriptional repressor SIN3 and the HDAC-1 histone deacetylase. Protein binding assays demonstrate that the LAZ3͞BCL6 BTB͞POZ domain directly interacts with SMRT in vitro. Furthermore, DNAbound LAZ3͞BCL6 recruits SMRT in vivo, and both overexpressed proteins completely colocalize in nuclear dots. Finally, overexpression of SMRT enhances the LAZ3͞BCL6-mediated repression. These results define SMRT as a corepressor of LAZ3͞BCL6 and suggest that LAZ3͞BCL6 and nuclear hormone receptors repress transcription through shared mechanisms involving SMRT recruitment and histone deacetylation.The LAZ3͞BCL6 (lymphoma-associated zinc finger 3͞B cell lymphomas 6) gene has been cloned by virtue of its frequent structural alteration in both diffuse large cell and follicular lymphomas (1-3). These alterations include translocations, small deletions, and point mutations. Most of them have been found in a genomic region, called the major translocation cluster, containing the first noncoding exon and the first downstream intron of the LAZ3͞BCL6 locus (4-8). It usually is proposed that such structural alterations lead to the deregulation of LAZ3͞BCL6 expression and, hence, contribute to lymphomagenesis (4, 7). The normal LAZ3͞BCL6 expression pattern suggests its implication in B cell differentiation and in the control of T cell-dependent immune response (9). Recent genetic experiments in mouse abrogating LAZ3͞BCL6 expression or leading to the expression of an inactive deleted version of this protein substantiate this hypothesis. Indeed, mice deficient for LAZ3͞BCL6 activity are devoid of germinal centers, present a Th2-type inflammatory disease and a defect in T cell-dependent antibody response (10, 11). Taken together, these results suggest that LAZ3͞BCL6-associated lymphomas may occur as a consequence of a deregulated LAZ3͞ BCL6 expression.The LAZ3͞BCL6 gene encodes a sequence-specific transcriptional repressor that harbors six C-terminal C2H2 krüp-pel-like zinc fingers. These zinc fingers are responsible for the sequence-specific DNA binding of the protein. At its Nterminal part, LAZ3͞BCL6 also contains an Ϸ130-aa conserved domain termed the BTB͞POZ (bric-à-brac tramtrack broad complex͞pox viruses and zinc fingers) domain (12,13). This domain has been identified in Ϸ40 proteins found in Metazoans and poxviruses (13). In LAZ3͞BCL6, the BTB͞ POZ domain mediates self-interaction an...

The labyrinth of the rodent placenta contains villi that are the site of nutrient exchange between mother and fetus. They are covered by three trophoblast cell types that separate the maternal blood sinusoids from fetal capillaries -a single mononuclear cell that is a subtype of trophoblast giant cell (sinusoidal or S-TGC) with endocrine function and two multinucleated syncytiotrophoblast layers, each resulting from cell-cell fusion, that function in nutrient transport. The developmental origins of these cell types have not previously been elucidated. We report here the discovery of cell-layer-restricted genes in the mid-gestation labyrinth (E12.5-14.5) including Ctsq in S-TGCs (also Hand1-positive), Syna in syncytiotrophoblast layer I (SynT-I), and Gcm1, Cebpa and Synb in syncytiotrophoblast layer II (SynT-II). These genes were also expressed in distinct layers in the chorion as early as E8.5, prior to villous formation. Specifically, Hand1 was expressed in apical cells lining maternal blood spaces (Ctsq is not expressed until E12.5), Syna in a layer immediately below, and Gcm1, Cebpa and Synb in basal cells in contact with the allantois. Cebpa and Synb were co-expressed with Gcm1 and were reduced in Gcm1 mutants. By contrast, Hand1 and Syna expression was unaltered in Gcm1 mutants, suggesting that Gcm1-positive cells are not required for the induction of the other chorion layers. These data indicate that the three differentiated trophoblast cell types in the labyrinth arise from distinct and autonomous precursors in the chorion that are patterned before morphogenesis begins.

Phosphorylation of transcription factors is regarded as a major mechanism to control their activity in regulation of gene expression. C/EBPp is a transcription factor that becomes activated after phosphorylation to induce genes involved in inflammation, acute-phase response, cytokine expression, cell growth, and differentiation. The chicken homolog NF-M collaborates with Myb and various kinase oncogenes in normal myeloid differentiation as well as in the leukemic transformation of myelomonocytic cells. Here, we examined the structure of NF-M and its mechanism of activation. We show that NF-M is a repressed transcription factor with concealed activation potential. Derepressed NF-M exhibits enhanced transcriptional efficacy in reporter assays. More importantly, NF-M activates resident chromatin-embedded, myelomonocyte-specific target genes, even in heterologous cell types such as fibroblasts or erythroblasts. We identified two regions within NF-M that act to repress trans-activation. Repression is abolished by deletion of these regions, activation of signal transduction kinases including v-erbB, polyoma middle T, ras and mil/raf, or point mutation of a critical phosphorylation site for MAP kinases. We provide evidence that phosphorylation plays a unique role to derepress rather than to enhance the trans-activation domain as a novel mechanism to regulate gene expression by NF-M/C/EBPp.