Herbal and natural medicines in the Latino community.
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Erectile Dysfunction↗
Biomedical subjects
Richard A Young
Publications and source records attributed to Richard A Young.
NOTCH1 directly regulates c-MYC and activates a feed-forward-loop transcriptional network promoting leukemic cell growth.
The NOTCH1 signaling pathway directly links extracellular signals with transcriptional responses in the cell nucleus and plays a critical role during T cell development and in the pathogenesis over 50% of human T cell lymphoblastic leukemia (T-ALL) cases. However, little is known about the transcriptional programs activated by NOTCH1. Using an integrative systems biology approach we show that NOTCH1 controls a feed-forward-loop transcriptional network that promotes cell growth. Inhibition of NOTCH1 signaling in T-ALL cells led to a reduction in cell size and elicited a gene expression signature dominated by down-regulated biosynthetic pathway genes. By integrating gene expression array and ChIP-on-chip data, we show that NOTCH1 directly activates multiple biosynthetic routes and induces c-MYC gene expression. Reverse engineering of regulatory networks from expression profiles showed that NOTCH1 and c-MYC govern two directly interconnected transcriptional programs containing common target genes that together regulate the growth of primary T-ALL cells. These results identify c-MYC as an essential mediator of NOTCH1 signaling and integrate NOTCH1 activation with oncogenic signaling pathways upstream of c-MYC.
Cell Line↗
Zebrafish promoter microarrays identify actively transcribed embryonic genes.
We have designed a zebrafish genomic microarray to identify DNA-protein interactions in the proximal promoter regions of over 11,000 zebrafish genes. Using these microarrays, together with chromatin immunoprecipitation with an antibody directed against tri-methylated lysine 4 of Histone H3, we demonstrate the feasibility of this method in zebrafish. This approach will allow investigators to determine the genomic binding locations of DNA interacting proteins during development and expedite the assembly of the genetic networks that regulate embryogenesis.
Animals↗
Activated signal transduction kinases frequently occupy target genes.
Cellular signal transduction pathways modify gene expression programs in response to changes in the environment, but the mechanisms by which these pathways regulate populations of genes under their control are not entirely understood. We present evidence that most mitogen-activated protein kinases and protein kinase A subunits become physically associated with the genes that they regulate in the yeast (Saccharomyces cerevisiae) genome. The ability to detect this interaction of signaling kinases with target genes can be used to more precisely and comprehensively map the regulatory circuitry that eukaryotic cells use to respond to their environment.
Adaptor Proteins, Signal Transducing↗
Core transcriptional regulatory circuitry in human hepatocytes.
We mapped the transcriptional regulatory circuitry for six master regulators in human hepatocytes using chromatin immunoprecipitation and high-resolution promoter microarrays. The results show that these regulators form a highly interconnected core circuitry, and reveal the local regulatory network motifs created by regulator-gene interactions. Autoregulation was a prominent theme among these regulators. We found that hepatocyte master regulators tend to bind promoter regions combinatorially and that the number of transcription factors bound to a promoter corresponds with observed gene expression. Our studies reveal portions of the core circuitry of human hepatocytes.
Gene Expression Regulation↗
Control of developmental regulators by Polycomb in human embryonic stem cells.
Polycomb group proteins are essential for early development in metazoans, but their contributions to human development are not well understood. We have mapped the Polycomb Repressive Complex 2 (PRC2) subunit SUZ12 across the entire nonrepeat portion of the genome in human embryonic stem (ES) cells. We found that SUZ12 is distributed across large portions of over two hundred genes encoding key developmental regulators. These genes are occupied by nucleosomes trimethylated at histone H3K27, are transcriptionally repressed, and contain some of the most highly conserved noncoding elements in the genome. We found that PRC2 target genes are preferentially activated during ES cell differentiation and that the ES cell regulators OCT4, SOX2, and NANOG cooccupy a significant subset of these genes. These results indicate that PRC2 occupies a special set of developmental genes in ES cells that must be repressed to maintain pluripotency and that are poised for activation during ES cell differentiation.
Animals↗
Polycomb complexes repress developmental regulators in murine embryonic stem cells.
The mechanisms by which embryonic stem (ES) cells self-renew while maintaining the ability to differentiate into virtually all adult cell types are not well understood. Polycomb group (PcG) proteins are transcriptional repressors that help to maintain cellular identity during metazoan development by epigenetic modification of chromatin structure. PcG proteins have essential roles in early embryonic development and have been implicated in ES cell pluripotency, but few of their target genes are known in mammals. Here we show that PcG proteins directly repress a large cohort of developmental regulators in murine ES cells, the expression of which would otherwise promote differentiation. Using genome-wide location analysis in murine ES cells, we found that the Polycomb repressive complexes PRC1 and PRC2 co-occupied 512 genes, many of which encode transcription factors with important roles in development. All of the co-occupied genes contained modified nucleosomes (trimethylated Lys 27 on histone H3). Consistent with a causal role in gene silencing in ES cells, PcG target genes were de-repressed in cells deficient for the PRC2 component Eed, and were preferentially activated on induction of differentiation. Our results indicate that dynamic repression of developmental pathways by Polycomb complexes may be required for maintaining ES cell pluripotency and plasticity during embryonic development.
Animals↗
Transcriptional regulatory networks downstream of TAL1/SCL in T-cell acute lymphoblastic leukemia.
Aberrant expression of 1 or more transcription factor oncogenes is a critical component of the molecular pathogenesis of human T-cell acute lymphoblastic leukemia (T-ALL); however, oncogenic transcriptional programs downstream of T-ALL oncogenes are mostly unknown. TAL1/SCL is a basic helix-loop-helix (bHLH) transcription factor oncogene aberrantly expressed in 60% of human T-ALLs. We used chromatin immunoprecipitation (ChIP) on chip to identify 71 direct transcriptional targets of TAL1/SCL. Promoters occupied by TAL1 were also frequently bound by the class I bHLH proteins E2A and HEB, suggesting that TAL1/E2A as well as TAL1/HEB heterodimers play a role in transformation of T-cell precursors. Using RNA interference, we demonstrated that TAL1 is required for the maintenance of the leukemic phenotype in Jurkat cells and showed that TAL1 binding can be associated with either repression or activation of genes whose promoters occupied by TAL1, E2A, and HEB. In addition, oligonucleotide microarray analysis of RNA from 47 primary T-ALL samples showed specific expression signatures involving TAL1 targets in TAL1-expressing compared with -nonexpressing human T-ALLs. Our results indicate that TAL1 may act as a bifunctional transcriptional regulator (activator and repressor) at the top of a complex regulatory network that disrupts normal T-cell homeostasis and contributes to leukemogenesis.
Basic Helix-Loop-Helix Proteins↗
Coordinated binding of NF-kappaB family members in the response of human cells to lipopolysaccharide.
The NF-kappaB family of transcription factors plays a critical role in numerous cellular processes, particularly the immune response. Our understanding of how the different NF-kappaB subunits act coordinately to regulate gene expression is based on a limited set of genes. We used genome-scale location analysis to identify targets of all five NF-kappaB proteins before and after stimulation of monocytic cells with bacterial lipopolysaccharide (LPS). In unstimulated cells, p50 and p52 bound to a large number of gene promoters that were also occupied by RNA polymerase II. After LPS stimulation, additional NF-kappaB subunits bound to these genes and to other genes. Genes that became bound by multiple NF-kappaB subunits were the most likely to show increases in RNA polymerase II occupancy and gene expression. This study identifies NF-kappaB target genes, reveals how the different NF-kappaB proteins coordinate their activity, and provides an initial map of the transcriptional regulatory network that underlies the host response to infection.
Genome, Human↗
Global and gene-specific analyses show distinct roles for Myod and Myog at a common set of promoters.
We used a combination of genome-wide and promoter-specific DNA binding and expression analyses to assess the functional roles of Myod and Myog in regulating the program of skeletal muscle gene expression. Our findings indicate that Myod and Myog have distinct regulatory roles at a similar set of target genes. At genes expressed throughout the program of myogenic differentiation, Myod can bind and recruit histone acetyltransferases. At early targets, Myod is sufficient for near full expression, whereas, at late expressed genes, Myod initiates regional histone modification but is not sufficient for gene expression. At these late genes, Myog does not bind efficiently without Myod; however, transcriptional activation requires the combined activity of Myod and Myog. Therefore, the role of Myog in mediating terminal differentiation is, in part, to enhance expression of a subset of genes previously initiated by Myod.
Acetylation↗
High-resolution computational models of genome binding events.
Direct physical information that describes where transcription factors, nucleosomes, modified histones, RNA polymerase II and other key proteins interact with the genome provides an invaluable mechanistic foundation for understanding complex programs of gene regulation. We present a method, joint binding deconvolution (JBD), which uses additional easily obtainable experimental data about chromatin immunoprecipitation (ChIP) to improve the spatial resolution of the transcription factor binding locations inferred from ChIP followed by DNA microarray hybridization (ChIP-Chip) data. Based on this probabilistic model of binding data, we further pursue improved spatial resolution by using sequence information. We produce positional priors that link ChIP-Chip data to sequence data by guiding motif discovery to inferred protein-DNA binding sites. We present results on the yeast transcription factors Gcn4 and Mig2 to demonstrate JBD's spatial resolution capabilities and show that positional priors allow computational discovery of the Mig2 motif when a standard approach fails.
Base Sequence↗
Evidence for an instructive mechanism of de novo methylation in cancer cells.
DNA methylation has a role in the regulation of gene expression during normal mammalian development but can also mediate epigenetic silencing of CpG island genes in cancer and other diseases. Many individual genes (including tumor suppressors) have been shown to undergo de novo methylation in specific tumor types, but the biological logic inherent in this process is not understood. To decipher this mechanism, we have adopted a new approach for detecting CpG island DNA methylation that can be used together with microarray technology. Genome-wide analysis by this technique demonstrated that tumor-specific methylated genes belong to distinct functional categories, have common sequence motifs in their promoters and are found in clusters on chromosomes. In addition, many are already repressed in normal cells. These results are consistent with the hypothesis that cancer-related de novo methylation may come about through an instructive mechanism.
Animals↗
Research participation, protected time, and research output by family physicians in family medicine residencies.
BACKGROUND AND OBJECTIVES: The Future of Family Medicine project concluded that research must become a greater part of the culture of the specialty. We examined the participation of family physician residency faculty in research, their protected time, and their research output and how these varied by program type. METHODS: This was a cross-sectional survey of all family medicine residency programs in the United States. The response rate was 66% (298/453). RESULTS: The majority of programs reported at least one family physician who participates in research, though the medical school-based (MSB) programs reported a higher total number of faculty than the community-based, medical school affiliated (MSA) programs (9.53 versus 2.72) and percentage of faculty (56% versus 37%). Substantially more MSB programs reported that they had at least one family physician with significant protected time for research (48% versus 7% for > 25% protected time) or any protected time (69% for MSB versus 45% for MSA). MSB programs and MSA programs reported similar success at producing at least one poster or paper for national meetings within the last 3 years (63% versus 41%) but not for published papers (86% versus 43%). CONCLUSIONS: We found that only about half of the family medicine residencies produced any nationally recognized research over a 3-year period and that this represents only a small improvement over the last 10 years. Our findings suggest that more support is needed if research is to become an integral part of the culture of family medicine.
Cross-Sectional Studies↗
The core centromere and Sgo1 establish a 50-kb cohesin-protected domain around centromeres during meiosis I.
The stepwise loss of cohesins, the complexes that hold sister chromatids together, is required for faithful meiotic chromosome segregation. Cohesins are removed from chromosome arms during meiosis I but are maintained around centromeres until meiosis II. Here we show that Sgo1, a protein required for protecting centromeric cohesins from removal during meiosis I, localizes to cohesin-associated regions (CARs) at the centromere and the 50-kb region surrounding it. Establishment of this Sgo1-binding domain requires the 120-base-pair (bp) core centromere, the kinetochore component Bub1, and the meiosis-specific factor Spo13. Interestingly, cohesins and the kinetochore proteins Iml3 and Chl4 are necessary for Sgo1 to associate with pericentric regions but less so for Sgo1 to associate with the core centromeric regions. Finally, we show that the 50-kb Sgo1-binding domain is the chromosomal region where cohesins are protected from removal during meiosis I. Our results identify the portions of chromosomes where cohesins are protected from removal during meiosis I and show that kinetochore components and cohesins themselves are required to establish this cohesin protective domain.
Cell Cycle Proteins↗
A hypothesis-based approach for identifying the binding specificity of regulatory proteins from chromatin immunoprecipitation data.
MOTIVATION: Genome-wide chromatin-immunoprecipitation (ChIP-chip) detects binding of transcriptional regulators to DNA in vivo at low resolution. Motif discovery algorithms can be used to discover sequence patterns in the bound regions that may be recognized by the immunoprecipitated protein. However, the discovered motifs often do not agree with the binding specificity of the protein, when it is known. RESULTS: We present a powerful approach to analyzing ChIP-chip data, called THEME, that tests hypotheses concerning the sequence specificity of a protein. Hypotheses are refined using constrained local optimization. Cross-validation provides a principled standard for selecting the optimal weighting of the hypothesis and the ChIP-chip data and for choosing the best refined hypothesis. We demonstrate how to derive hypotheses for proteins from 36 domain families. Using THEME together with these hypotheses, we analyze ChIP-chip datasets for 14 human and mouse proteins. In all the cases the identified motifs are consistent with the published data with regard to the binding specificity of the proteins.
Algorithms↗
Core transcriptional regulatory circuitry in human embryonic stem cells.
The transcription factors OCT4, SOX2, and NANOG have essential roles in early development and are required for the propagation of undifferentiated embryonic stem (ES) cells in culture. To gain insights into transcriptional regulation of human ES cells, we have identified OCT4, SOX2, and NANOG target genes using genome-scale location analysis. We found, surprisingly, that OCT4, SOX2, and NANOG co-occupy a substantial portion of their target genes. These target genes frequently encode transcription factors, many of which are developmentally important homeodomain proteins. Our data also indicate that OCT4, SOX2, and NANOG collaborate to form regulatory circuitry consisting of autoregulatory and feedforward loops. These results provide new insights into the transcriptional regulation of stem cells and reveal how OCT4, SOX2, and NANOG contribute to pluripotency and self-renewal.
Animals↗
Genome-wide map of nucleosome acetylation and methylation in yeast.
Eukaryotic genomes are packaged into nucleosomes whose position and chemical modification state can profoundly influence regulation of gene expression. We profiled nucleosome modifications across the yeast genome using chromatin immunoprecipitation coupled with DNA microarrays to produce high-resolution genome-wide maps of histone acetylation and methylation. These maps take into account changes in nucleosome occupancy at actively transcribed genes and, in doing so, revise previous assessments of the modifications associated with gene expression. Both acetylation and methylation of histones are associated with transcriptional activity, but the former occurs predominantly at the beginning of genes, whereas the latter can occur throughout transcribed regions. Most notably, specific methylation events are associated with the beginning, middle, and end of actively transcribed genes. These maps provide the foundation for further understanding the roles of chromatin in gene expression and genome maintenance.
Acetylation↗
Binding of pRB to the PHD protein RBP2 promotes cellular differentiation.
pRB can enforce a G1 block by repressing E2F-responsive promoters. It also coactivates certain non-E2F transcription factors and promotes differentiation. Some pRB variants activate transcription and promote differentiation despite impaired E2F binding and transcriptional repression capabilities. We identified RBP2 in a screen for proteins that bind to such pRB variants. RBP2 resembles other chromatin-associated transcriptional regulators and RBP2 binding tracked with pRB's ability to activate transcription and promote differentiation. RBP2 and pRB colocalize and pRB/RBP2 complexes were detected in chromatin isolated from differentiating cells. RBP2 siRNA phenocopied restoration of pRB function in coactivation and differentiation assays, suggesting that pRB prevents RBP2 from repressing genes required for differentiation. In addition, two bromodomain-containing proteins were identified as RBP2 targets that are transcriptionally activated by pRB in an RBP2-dependent manner. Our results suggest that promotion of differentiation by pRB involves neutralization of free RBP2 and transcriptional activation of RBP2 targets linked to euchromatin maintenance.
Animals↗