Cell-Type-Specific Activation and Repression of PU.1 by a Complex of Discrete, Functionally Specialized cis-Regulatory Elements

The transcription factor PU.1 is critical for multiple hematopoietic lineages, but different leukocyte types require strictly distinct patterns of PU.1 regulation. PU.1 is required early for T-cell lineage development but then must be repressed by a stage-specific mechanism correlated with commitment. Other lineages require steady, low expression or upregulation. Until now, only the promoter plus a distal upstream regulatory element (URE) could be invoked to explain nearly all Sfpi1 (PU.1) activation and repression, including bifunctional effects of Runx1. However, the URE is dispensable for most Sfpi1 downregulation in early T cells, and we show that it retains enhancer activity in immature T-lineage cells even where endogenous Sfpi1 is repressed. We now present evidence for another complex of conserved noncoding elements that mediate discrete, cell-type-specific regulatory features of Sfpi1, including a myeloid cell-specific activating element and a separate, pro-T-cell-specific silencer element. These elements yield opposite, cell-type-specific responses to Runx1. T-cell-specific repression requires Runx1 acting through multiple nonconsensus sites in the silencer core. These newly characterized sites recruit Runx1 binding in early T cells in vivo and define a functionally specific scaffold for dose-dependent, Runx-mediated repression

A gene regulatory network armature for T lymphocyte specification

Choice of a T lymphoid fate by hematopoietic progenitor cells depends on sustained Notch–Delta signaling combined with tightly regulated activities of multiple transcription factors. To dissect the regulatory network connections that mediate this process, we have used high-resolution analysis of regulatory gene expression trajectories from the beginning to the end of specification, tests of the short-term Notch dependence of these gene expression changes, and analyses of the effects of overexpression of two essential transcription factors, namely PU.1 and GATA-3. Quantitative expression measurements of >50 transcription factor and marker genes have been used to derive the principal components of regulatory change through which T cell precursors progress from primitive multipotency to T lineage commitment. Our analyses reveal separate contributions of Notch signaling, GATA-3 activity, and down-regulation of PU.1. Using BioTapestry (www.BioTapestry.org), the results have been assembled into a draft gene regulatory network for the specification of T cell precursors and the choice of T as opposed to myeloid/dendritic or mast-cell fates. This network also accommodates effects of E proteins and mutual repression circuits of Gfi1 against Egr-2 and of TCF-1 against PU.1 as proposed elsewhere, but requires additional functions that remain unidentified. Distinctive features of this network structure include the intense dose dependence of GATA-3 effects, the gene-specific modulation of PU.1 activity based on Notch activity, the lack of direct opposition between PU.1 and GATA-3, and the need for a distinct, late-acting repressive function or functions to extinguish stem and progenitor-derived regulatory gene expression

Characteristics of contralateral carcinomas in patients with differentiated thyroid cancer larger than 1 cm

Purpose: Traditionally, total thyroidectomy has been advocated for patients with tumors larger than 1 cm. However, according to the ATA and NCCN guidelines (2015, USA), patients with tumors up to 4 cm are now eligible for lobectomy. A rationale for adhering to total thyroidectomy might be the presence of contralateral carcinomas. The purpose of this study was to describe the characteristics of contralateral carcinomas in patients with differentiated thyroid cancer (DTC) larger than 1 cm. Methods: A retrospective study was performed including patients from 17 centers in 5 countries. Adults diagnosed with DTC stage T1b-T3 N0-1a M0 who all underwent a total thyroidectomy were included. The primary endpoint was the presence of a contralateral carcinoma. Results: A total of 1

Characteristics of contralateral carcinomas in patients with differentiated thyroid cancer larger than 1 cm

textabstractPurpose: Traditionally, total thyroidectomy has been advocated for patients with tumors larger than 1 cm. However, according to the ATA and NCCN guidelines (2015, USA), patients with tumors up to 4 cm are now eligible for lobectomy. A rationale for adhering to total thyroidectomy might be the presence of contralateral carcinomas. The purpose of this study was to describe the characteristics of contralateral carcinomas in patients with differentiated thyroid cancer (DTC) larger than 1 cm. Methods: A retrospective study was performed including patients from 17 centers in 5 countries. Adults diagnosed with DTC stage T1b-T3 N0-1a M0 who all underwent a total thyroidectomy were included. The primary endpoint was the presence of a contralateral carcinoma. Results: A total of 1313 patients were included, of whom 426 (32 %) had a contralateral carcinoma. The contralateral carcinomas consisted of 288 (67 %) papillary thyroid carcinomas (PTC), 124 (30 %) follicular variant of a papillary thyroid carcinoma (FvPTC), 5 (1 %) follicular thyroid carcinomas (FTC), and 3 (1 %) Hürthle cell carcinomas (HTC). Ipsilateral multifocality was strongly associated with the presence of contralateral carcinomas (OR 2.62). Of all contralateral carcinomas, 82 % were ≤10 mm and of those 99 % were PTC or FvPTC. Even if the primary tumor was a FTC or HTC, the contralateral carcinoma was (Fv)PTC in 92 % of cases. Conclusions: This international multicenter study performed on patients with DTC larger than 1 cm shows that contralateral carcinomas occur in one third of patients and, independently of primary tumor subtype, predominantly consist of microPTC

Magic-factor 1, a partial agonist of Met, induces muscle hypertrophy by protecting myogenic progenitors from apoptosis.

Hepatocyte Growth Factor (HGF) is a pleiotropic cytokine of mesenchymal origin that mediates a characteristic array of biological activities including cell proliferation, survival, motility and morphogenesis. Its high affinity receptor, the tyrosine kinase Met, is expressed by a wide range of tissues and can be activated by either paracrine or autocrine stimulation. Adult myogenic precursor cells, the so called satellite cells, express both HGF and Met. Following muscle injury, autocrine HGF-Met stimulation plays a key role in promoting activation and early division of satellite cells, but is shut off in a second phase to allow myogenic differentiation. In culture, HGF stimulation promotes proliferation of muscle precursors thereby inhibiting their differentiation

Investigating the Transcriptional Mechanisms Controlling Sfpi1, a Critical Regulatory Node Within Multiple Lineage Specifying Subcircuits of the Hematopoietic Gene Regulatory Network

The Sfpi1 gene encodes PU.1, a critical transcription factor in multiple hematopoietic lineages. PU.1 expression is upregulated as hematopoietic stem cells become granulocyte-macrophage progenitors. In contrast, Sfpi1 must be silenced after progenitors undergo T-lineage specification. If unrestrained in early T-lineage cells, PU.1 can both block developmental progress and induce diversion to a myeloid fate. When PU.1 expression is not sufficiently increased or maintained in myeloid lineage cells, myeloid hyperproliferation and cancer can result. In mouse DN thymocytes, PU.1 mRNA begins at high levels in early T-cell progenitors, but drops about fivefold as cells enter the T-cell program (DN2) and then falls tenfold further as the cells reach T-lineage commitment (DN3). This implies operation of a stage-specific repression mechanism correlated with commitment. Only one major cis-regulatory element has previously been described for Sfpi1, which is a compound conserved region around -14 kb that is thought to mediate activation as well as some repression. However, it cannot account for all PU.1 regulation in early T-lineage cells nor in myeloid cells. In particular, that -14 kb element can show strong enhancer activity in an immature T-cell line in which the endogenous Sfpi1 gene is profoundly repressed. Additionally, absence of the -14 kb element does not abolish PU.1 expression in myeloid lineages. We now present evidence for another complex of conserved noncoding elements that appear to mediate several cell-type-specific regulatory features, including cell-type-specific repression in early T-cells. We describe fine mapping of a T-cell specific bipartite silencer and show that the T lineage specific repressive activity requires Runx1. We also describe additional regulatory complexes that may contribute to lineage specific regulation of PU.1 in early hematopoietic progenitors, including a myeloid specific enhancer. We provide evidence of lineage restricted occupancy of these additional regulatory elements and show that the novel enhancer elements are additional sites of PU.1 auto regulation