Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Chromatin loop”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,117 records · Page 62Linked to original sources
PubMed2005

The stem-loop binding protein regulates translation of histone mRNA during mammalian oogenesis.

Although messenger RNAs encoding the histone proteins are among the most abundant in mammalian oocytes, the mechanism regulating their translation has not been identified. The stem-loop binding protein (SLBP) binds to a highly conserved sequence in the 3'-untranslated region (utr) of the non-polyadenylated histone mRNAs in somatic cells and mediates their stabilization and translation. We previously showed that SLBP, which is expressed only during S-phase of proliferating cells, is expressed in growing oocytes at G2 of the cell cycle and accumulates substantially during meiotic maturation. We report here that elevating the amount of SLBP in immature (G2) oocytes is sufficient to increase translation of a reporter mRNA bearing the histone 3'-utr and endogenous histone synthesis and that this effect is not mediated through increased stability of the encoding mRNAs. We further report that translation of the reporter mRNA increases dramatically during meiotic maturation coincident with the accumulation of SLBP. Conversely, when SLBP accumulation during maturation is prevented using RNA interference, both translation of the reporter mRNA and synthesis of endogenous histones are significantly reduced. This effect is not mediated by a loss of the encoding mRNAs. Moreover, following fertilization, SLBP-depleted oocytes also show a significant decrease in pronuclear size and in the amount of acetylated histone detectable on the chromatin. These results demonstrate that histone synthesis in immature and maturing oocytes is governed by a translational control mechanism that is directly regulated by changes in the amount of SLBP.

3' Untranslated Regions↗
PubMed2002

p8-deficient fibroblasts grow more rapidly and are more resistant to adriamycin-induced apoptosis.

p8 is a stress-induced DNA-binding protein, biochemically related to the architectural chromatin binding HMG protein family and whose function is presently unknown. We obtained fibroblast from mice lacking p8 and found that p8 is involved in cell growth regulation and in apoptosis. p8(-/-) mouse embryonic fibroblasts (MEFs) grow more rapidly than p8(+/+) MEFs. This might be explained by the higher intracellular level and activity of the Cdk2 and Cdk4 observed in p8(-/-) MEFs, which in turn may result, at least in part, from the concomitant decrease observed in the amount of cyclin-dependent kinase inhibitor p27. We also report that p8 mRNA expression is strongly activated in fibroblasts after cell growth arrest induced by serum deprivation or confluence. As expected, MEFs expressing p8 arrest their growth more rapidly after serum deprivation than MEFs lacking p8, which strongly suggests that p8 over-expression is implicated in cell growth arrest. On the other hand, p8(+/+) MEFs are more sensitive than p8(-/-) MEFs to the apoptosis induced by adriamycin treatment. p53 might be involved, as p8 expression increases its intracellular amount and trans-activation capacity. Finally, demonstration that p53 is a negative trans-activator of p8 suggests the presence of a complex autoregulatory loop. In conclusion, p8 is a cell growth inhibitor that facilitates apoptosis induced in fibroblasts by DNA damage.

Animals↗
PubMed2026

Dual Mechanisms Underlie the Repression of Repressor Element 1-Silencing Transcription Factor Expression in Lung Neuroendocrine Carcinoma Cells.

Recent advances in genetic analysis have led to further subtyping of small-cell lung carcinoma (SCLC). The major subtypes of SCLC are the achaete-scute family bHLH transcription factor 1 (ASCL1)-predominant (SCLC-A), neuronal differentiation 1 (NEUROD1)-predominant (SCLC-N), and POU class 2 homeobox 3 (POU2F3) (SCLC-P) subtypes. SCLC-A and SCLC-N tumors express chromogranin A (CHGA) and synaptophysin (SYP), but SCLC-P tumors do not. Large-cell neuroendocrine carcinoma, another type of neuroendocrine carcinoma (NEC), also frequently expresses CHGA and SYP. Because CHGA and SYP expression is controlled by a transrepressor, repressor element 1-silencing transcription factor (REST), the mechanisms underlying REST suppression in NEC were investigated, with a focus on miRNAs and epigenetics, to determine the causes of the differences in the expression of CHGA and SYP between SCLC-A/N and SCLC-P cells. The results showed that miR-375-3p, which was induced by ASCL1 and NEUROD1, repressed REST expression by binding to the 3'-untranslated region of REST mRNA. Bisulfite sequencing and experiments using a DNA methyltransferase inhibitor, a histone deacetylase inhibitor, and chromatin immunoprecipitation-based quantitative PCR revealed that promoter/enhancer hypermethylation and histone deacetylation causes REST gene inactivation in SCLC-A/N. These phenomena were also observed in a large-cell neuroendocrine carcinoma cell line that expressed high levels of ASCL1 and NEUROD1. Taken together, these findings suggest that NEC has dual repressive effects on REST expression, resulting in strict regulation of the expression of CHGA, SYP, and other REST-controlled neuronal/neuroendocrine-specific genes.

Humans↗
PubMed2000

Interaction of MyoD family proteins with enhancers of acetylcholine receptor subunit genes in vivo.

The myogenic determination factors (MDFs) are transcriptional activators that target E boxes in many muscle-specific promoters, including those of the genes coding for the subunits of the acetylcholine receptor. It is not known, however, if in vivo a given E box in a transcriptionally active gene is occupied, either uniquely by one MDF or randomly by all MDFs. We have analyzed expression of MDF and acetylcholine receptor subunits in cultured mouse muscle cells and, using chromatin immunoprecipitation, have determined which individual MDFs reside at promoters of several receptor subunit genes. We find that before fusion, C2C12 cells express myf-5, MyoD, and myogenin, all of which take up residence at promoters of all subunits except epsilon. At this stage, herculin is present in limited amounts and is detected mainly at the gamma and delta subunit genes. On myotube formation, herculin reaches high levels; concomitantly, the epsilon subunit gene becomes a common MDF target and begins to be expressed. In general, any MDF protein that is expressed also is present on transcriptionally active receptor genes; transcriptional activity of target genes correlates with occupancy by MDF, in particular, herculin.

Animals↗
PubMed1986

Cohabitation of scaffold binding regions with upstream/enhancer elements of three developmentally regulated genes of D. melanogaster.

We find DNA fragments attached to the nuclear scaffold (SARs) both 5' and 3' of three Drosophila genes, defining looped domains ranging from 4.5 to 13 kb. For the two-promoter-containing gene Adh (alcohol dehydrogenase), we find two upstream and two downstream SARs. For Sgs-4, the 5' SAR covers 866 bp immediately upstream of the transcript, and in the case of fushi tarazu, the 5' SAR is found on a small fragment 4.8 kb upstream of the start of transcription. These four upstream scaffold-attached fragments comap with enhancer-like regulatory sequences. Sequence analysis of five upstream SARs reveals clusters of sequences closely related to the cleavage consensus of topoisomerase II, several copies of a specific 10 bp A-rich sequence (AATAAATCAAA), and another 10 bp T-rich stretch.

Alcohol Dehydrogenase↗
PubMed2003

Role of CCAAT/enhancer-binding protein alpha (C/EBPalpha) in activation of the Kaposi's sarcoma-associated herpesvirus (KSHV) lytic-cycle replication-associated protein (RAP) promoter in cooperation with the KSHV replication and transcription activator (RTA) and RAP.

The Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded replication-associated protein (RAP, or K8) has been shown to induce both CCAAT/enhancer binding protein alpha (C/EBPalpha) and p21(CIP-1) expression, resulting in G(0)/G(1) cell cycle arrest during the lytic cycle. RAP and C/EBPalpha are also known to interact strongly both in vitro and in lytically infected cells. We recognized two potential consensus C/EBP binding sites in the RAP promoter and performed electrophoretic mobility shift assay (EMSA) analysis with in vitro-translated C/EBPalpha; this analysis showed that one of these sites has a very high affinity for C/EBPalpha. Luciferase (LUC) assays performed with a target RAP promoter-LUC reporter gene confirmed that C/EBPalpha can transcriptionally activate the RAP promoter up to 50-fold. Although RAP had no effect on its own promoter by itself, the addition of RAP and C/EBPalpha together resulted in a threefold increase in activity over that obtained with C/EBPalpha alone. Importantly, the introduction of exogenous Flag-tagged C/EBPalpha triggered RAP expression in BCBL-1 cells latently infected with KSHV, as detected by both reverse transcription-PCR and double-label immunofluorescence assay analyses, suggesting the presence of a self-reinforcing loop with C/EBPalpha and RAP activating each other. The RAP promoter can also be activated 50- to 120-fold by the KSHV lytic-cycle-triggering protein known as replication and transcription activator (RTA). C/EBPalpha and RTA together cooperated to elevate RAP promoter activity four- to sixfold more than either alone. Furthermore, the addition of RAP, C/EBPalpha, and RTA in LUC reporter cotransfection assays resulted in 7- to 15-fold more activation than that seen with either C/EBPalpha or RTA alone. Site-specific mutational analysis of the RAP promoter showed that the strong C/EBP binding site is crucial for C/EBPalpha-mediated transactivation of the RAP promoter. However, the C/EBP binding site also overlaps the previously reported 16-bp RTA-responsive element (RRE), and the same mutation also both reduced RTA-mediated transactivation and abolished the cooperativity between C/EBPalpha and RTA. Furthermore, in vitro-translated RTA, although capable of binding directly to the polyadenylated nuclear RNA (PAN) RRE motif, failed to bind to the RAP RRE and interfered with RRE-bound C/EBPalpha in EMSA experiments. Partial RTA responsiveness but no cooperativity could be transferred to a heterologous promoter containing added consensus C/EBP binding sites. A chromatin immunoprecipitation assay showed that all three proteins associated specifically with RAP promoter DNA in vivo and that, when C/EBPalpha was removed from a tetradecanoyl phorbol acetate-treated JSC-1 primary effusion lymphoma cell lysate, the levels of association of RTA and RAP with the RAP promoter were reduced 3- and 13-fold, respectively. Finally, RTA also proved to physically interact with both C/EBPalpha and RAP, as assayed both in vitro and by immunoprecipitation. Binding to C/EBPalpha occurred within the N-terminal DNA binding domain of RTA, and deletion of a 17-amino-acid basic motif of RTA abolished both the C/EBPalpha and DNA binding activities as well as all RTA transactivation and the cooperativity with C/EBPalpha. Therefore, we suggest that RTA transactivation of the RAP RRE is mediated by an interaction with DNA-bound C/EBPalpha but that full activity requires more than just the core C/EBP binding site.

Amino Acid Sequence↗
PubMed2002

Transcription factor occupancy of the insulin gene in vivo. Evidence for direct regulation by Nkx2.2.

Consensus-binding sites for many transcription factors are relatively non-selective and found at high frequency within the genome. This raises the possibility that factors that are capable of binding to a cis-acting element in vitro and regulating transcription from a transiently transfected plasmid, which would not have higher order chromatin structure, may not occupy this site within the endogenous gene. Closed chromatin structure and competition from another DNA-binding protein with similar nucleotide specificity are two possible mechanisms by which a transcription factor may be excluded from a potential binding site in vivo. Multiple transcription factors, including Pdx-1, BETA-2, and Pax6, have been implicated in expression of the insulin gene in pancreatic beta cells. In this study, the chromatin immunoprecipitation assay has been used to show that these factors do, in fact, bind to insulin control region sequences in intact beta cells. In addition, another key islet-enriched transcription factor, Nkx2.2, was found to occupy this region using the chromatin immunoprecipitation assay. In vitro DNA-binding and transient transfection assays defined how Nkx2.2 affected insulin gene expression. Pdx-1 was also shown to bind within a region of the endogenous islet amyloid polypeptide, pax-4, and glucokinase genes that were associated with control in vitro. Because Pdx-1 does not regulate gene transcription in isolation, these sequences were examined for occupancy by the other insulin transcriptional regulators. BETA-2, Pax6, and Nkx2.2 were also found to bind to amyloid polypeptide, glucokinase, and pax-4 control sequences in vivo. These studies reveal the broad application of the Pdx-1, BETA-2, Pax6, and Nkx2.2 transcription factors in regulating expression of genes selectively expressed in islet beta cells.

Amyloid↗
PubMed1999

Defects in components of the proteasome enhance transcriptional silencing at fission yeast centromeres and impair chromosome segregation.

Fission yeast centromeres are transcriptionally silent and form a heterochromatin-like structure essential for normal centromere function; this appears analogous to heterochromatin and position effect variegation in other eukaryotes. Conditional mutations in three genes designated cep (centromere enhancer of position effect) were found to enhance transcriptional silencing within centromeres. Cloning of the cep1(+) and cep2(+) genes by functional complementation revealed that they are identical to the previously described genes pad1(+) and mts2(+), respectively, which both encode subunits of the proteasome 19S cap. Like Mts2 and Mts4, epitope-tagged Cep1/Pad1 localizes to or near the nuclear envelope throughout the cell cycle. The cep mutants display a range of phenotypes depending on the temperature. Silencing within the central domain of centromeres is increased at 36 degrees C. This suggests that the proteasome is involved in regulating silencing and thus centromeric chromatin architecture, possibly by lowering the level of some chromatin-associated protein by ubiquitin-dependent degradation. This is the first report of defective proteasome function affecting heterochromatin-mediated transcriptional silencing. At 36 and 32 degrees C, the cep mutants lose chromosomes at an elevated rate, and at 18 degrees C, the mutants are cryosensitive for growth. Cytological analysis at 18 degrees C revealed a defect in sister chromatid separation while other mitotic events occurred normally, indicating that cep mutations might interfere specifically with the degradation of inhibitor(s) of sister chromatid separation. These observations suggest that 19S subunits confer a level of substrate specificity on the proteasome and raise the possibility of a link between components involved in centromere architecture and sister chromatid cohesion.

Basic Helix-Loop-Helix Leucine Zipper Transcriptio↗
PubMed1997

Postprocedural Pap Smears: a LEEP of faith?

Loop electrosurgical excision procedure (LEEP) is gaining in popularity in the United States as an outpatient alternative to the diagnosis, and potentially the treatment, of cervical intraepithelial neoplasia (CIN). LEEP is fast, simple, performed under local anesthesia, readily learned, and without significant morbidity. As cytopathologists and cytotechnologists, immediate cytologic evaluation of cervico-vaginal smears following LEEP is not the routine; however, there are very specific artifacts, most of which are related to the transfer of thermal energy, which result from the procedure. It is important to recognize these cytomorphologic features for accurate interpretation. The indications and contraindications for LEEP are similar to those for other ablative or excisional procedures. There appears to sacrifice in the efficacy of diagnosing and treating CIN by this method. Factors predictive of disease clearance are as confounding as they are for any other cone procedure. At the University of Iowa Hospital and Clinics (UIHC), immediate post-LEEP endocervical brush (PLEB) is often performed as a method of assessing the endocervical canal for residual disease or skip lesions. The most common cytomorphologic features observed are: "taffy-pulled" nuclei in elongated endocervical cells; cell aggregates with coalesced cytoplasm; hockey stick nuclei; notched and enlarged nuclei; and, smudgy chromatin. The difficulties or most frequent diagnostic dilemmas in interpreting these smears initially include abundant blood and smudgy chromatin, often tempting an interpretation of "unsatisfactory". However, careful study reveals that these changes are related to the nature of the procedure and reproducible. Recognition and familiarization of these features enables more accurate interpretation of PLEB cytology. The significance of abnormal PLEB, with regard to disease clearance, is still uncertain.

Artifacts↗
PubMed1998

Nucleosomes inhibit phagocytosis of apoptotic thymocytes by peritoneal macrophages from MRL+/+ lupus-prone mice.

The nucleosome, the basic structure of chromatin and normal product of cell apoptosis, plays a pivotal role both in the induction and the pathogenesis of systemic lupus erythematosus (SLE). Nucleosomes have been found to circulate at high levels in patients with SLE and apoptosis of lymphoid cells is increased during human and murine lupus. In this study, we examined the presence of possible defects in clearance mechanisms of apoptotic cells in murine lupus, and questioned further whether nucleosomes could compromise this phagocytic process. There did not appear to be any intrinsic functional defect of macrophages from young MRL+/+ lupus-prone mice to recognize and phagocytose apoptotic thymocytes. Nucleosomes, as a mimic of increased cell apoptotsis in vivo, induced a strong, dose-dependent, inhibition of phagocytosis of apoptotic thymocytes by young, pre-autoimmune, macrophages of MRL+/+ mice, whereas macrophages of non-autoimmune C3H mice only exhibited a trend to inhibition. The nucleosome-elicited inhibitory effect persisted during the development of the autoimmune response and appeared to be specific for the molecular mechanisms involved in macrophage phagocytosis of apoptotic cells. Our data suggest that nucleosome elicited inhibition of phagocytosis of apoptotic cells by MRL+/+ macrophages before the onset of the autoimmune response contribute, in a positive loop, to sustain and/or augment the levels of circulating (and potentially immunogenic) nucleosomes in lupus.

Age Factors↗
PubMed1999

Two MAD tails: what the recent knockouts of Mad1 and Mxi1 tell us about the MYC/MAX/MAD network.

Members of the MAD/MXI protein family heterodimerize with MAX and repress transcription by recruiting a chromatin-modifying co-repressor complex to specific DNA target genes. Repression mediated by MAD is thought to antagonize the transcriptional activation and proliferation-promoting functions of MYC-MAX heterodimers. Because they are induced during differentiation, it has been suggested that MAD proteins act to limit cell proliferation during terminal differentiation. There is also controversial evidence that these proteins may function as tumor suppressors. Recently, targeted gene deletions of two members of this gene family, Mad1 and Mxi1, have been carried out in mice. Although these animals display what appear to be quite different phenotypes, further analysis supports the view that both these proteins function in cell-cycle exit during terminal differentiation, and that at least MXI1 can act as a tumor suppressor.

Animals↗
PubMed1987

Genomic organization and developmental fate of adjacent repeated sequences in a foldback DNA clone of Tetrahymena thermophila.

DNA sequence elimination and rearrangement occurs during the development of somatic cell lineages of eukaryotes and was first discovered over a century ago. However, the significance and mechanism of chromatin elimination are not understood. DNA elimination also occurs during the development of the somatic macronucleus from the germinal micronucleus in unicellular ciliated protozoa such as Tetrahymena thermophila. In this study foldback DNA from the micronucleus was used as a probe to isolate ten clones. All of those tested (4/4) contained sequences that were repetitive in the micronucleus and rearranged in the macronucleus. The presence of inverted repeated sequences was clearly demonstrated in one of them by electron microscopy. DNA sequence analysis showed that the left portion of this clone contains three tandem, directly repeated copies of a 340-bp sequence, a 120-bp portion of which appears in inverted orientation at a 1.6-kb distance. This clone, pTtFB1, was subjected to a detailed analysis of its developmental fate. Subregions were subcloned and used as probes against Southern blots of micronuclear and macronuclear DNA. We found that all subregions defined repeated sequence families in the micronuclear genome. A minimum of four different families was defined, two of which are retained in the macronucleus and two of which are completely eliminated. The inverted repeat family is retained with little rearrangement. Two of the families, defined by subregions that do not contain parts of the inverted repeat, one in the "loop" and one in the "right flanking region," are totally eliminated during macronuclear development--and contain open reading frames. A fourth family occurs in the "loop" region and is rearranged extensively during development. The two gene families that are eliminated are stable in the micronuclear genome but are not clustered together as evidenced by experiments in which DNAs from nullisomic strains are used to map family members to specific micronuclear chromosomes. The inverted repeat family is also stable in the micronuclear genome and is dispersed among several chromosomes. The significance of retained inverted repeats to the process of elimination is discussed.

Animals↗
PubMed2006

Target hub proteins serve as master regulators of development in yeast.

To understand the organization of the transcriptional networks that govern cell differentiation, we have investigated the transcriptional circuitry controlling pseudohyphal development in Saccharomyces cerevisiae. The binding targets of Ste12, Tec1, Sok2, Phd1, Mga1, and Flo8 were globally mapped across the yeast genome. The factors and their targets form a complex binding network, containing patterns characteristic of autoregulation, feedback and feed-forward loops, and cross-talk. Combinatorial binding to intergenic regions was commonly observed, which allowed for the identification of a novel binding association between Mga1 and Flo8, in which Mga1 requires Flo8 for binding to promoter regions. Further analysis of the network showed that the promoters of MGA1 and PHD1 were bound by all of the factors used in this study, identifying them as key target hubs. Overexpression of either of these two proteins specifically induced pseudohyphal growth under noninducing conditions, highlighting them as master regulators of the system. Our results indicate that target hubs can serve as master regulators whose activity is sufficient for the induction of complex developmental responses and therefore represent important regulatory nodes in biological networks.

Blotting, Western↗
PubMed1987

The formation, identification, and significance of DNA-protein cross-links in mammalian cells.

DNA-protein cross-links (DPC) are formed by a variety of radiations and chemicals which act via free radical formation. Covalency is inferred from the resistance of the cross-links to harsh treatments. In mammalian cells, a background of DPC (6000 per V79 cell) may result from normal associations of chromosomal loops with the nuclear protein matrix. After ionizing radiation, the elevated level of DPC (150 per Gy per V79 cell) are enriched in actively transcribing DNA and in a subset of proteins of the nuclear matrix. DPC formation is reduced by hydroxyl radical scavengers, by oxygen, and by hypertonic medium and is enhanced by hypotonic medium and by removal of intracellular glutathione. DPC are repaired more slowly than single-strand breaks and not at all when formed during metaphase. During the postirradiation period, changes in the sequence composition of the DNA of residual DPC are consistent with the preferential repair of DPC in actively expressed genes. Excision repair mechanisms have been proposed. Unrepaired DPC may block normal functions of the nuclear matrix, such as replication and transcription.

Animals↗
PubMed2006

Proteomic discovery of Max as a novel interacting partner of C/EBPalpha: a Myc/Max/Mad link.

The transcription factor CCAAT/enhancer binding protein a (C/EBPalpha) is important in the regulation of granulopoiesis and is disrupted in human acute myeloid leukemia. In the present study, we sought to identify novel C/EBPalpha interacting proteins in vivo through immunoprecipitation using mass spectrometry-based proteomic techniques. We identified Max, a heterodimeric partner of Myc, as one of the interacting proteins of C/EBPalpha in our screen. We confirmed the in vivo interaction of C/EBPalpha with Max and showed that this interaction involves the basic region of C/EBPalpha. Endogenous C/EBPalpha and Max, but not Myc and Max, colocalize in intranuclear structures during granulocytic differentiation of myeloid U937 cells. Max enhanced the transactivation capacity of C/EBPalpha on a minimal promoter. A chromatin immunoprecipitation assay revealed occupancy of the human C/EBPalpha promoter in vivo by Max and Myc under cellular settings and by C/EBPalpha and Max under retinoic acid induced granulocytic differentiation. Interestingly, enforced expression of Max and C/EBPalpha results in granulocytic differentiation of the human hematopoietic CD34(+) cells, as evidenced by CD11b, CD15 and granulocyte colony-stimulating factor receptor expression. Silencing of Max by short hairpin RNA in CD34(+) and U937 cells strongly reduced the differentiation-inducing potential of C/EBPalpha, indicating the importance of C/EBPalpha-Max in myeloid progenitor differentiation. Taken together, our data reveal Max as a novel co-activator of C/EBPalpha functions, thereby suggesting a possible link between C/EBPalpha and Myc-Max-Mad network.

Basic Helix-Loop-Helix Leucine Zipper Transcriptio↗
PubMed2006

CTCF mediates interchromosomal colocalization between Igf2/H19 and Wsb1/Nf1.

Gene transcription may be regulated by remote enhancer or insulator regions through chromosome looping. Using a modification of chromosome conformation capture (3C) and fluorescence in situ hybridization, we found that one allele of the insulin-like growth factor 2 (Igf2)/H19 imprinting control region (ICR) on chromosome 7 colocalized with one allele of Wsb1/Nf1 on chromosome 11. Omission of CCCTC-binding factor (CTCF) or deletion of the maternal ICR abrogated this association and altered Wsb1/Nf1 gene expression. These findings demonstrate that CTCF mediates an interchromosomal association, perhaps by directing distant DNA segments to a common transcription factory, and the data provide a model for long-range allele-specific associations between gene regions on different chromosomes that suggest a framework for DNA recombination and RNA trans-splicing.

Alleles↗
PubMed2006

ChREBP binding to fatty acid synthase and L-type pyruvate kinase genes is stimulated by glucose in pancreatic beta-cells.

Pancreatic beta-cell dysfunction is central to the pathogenesis of type 2 diabetes and may involve secretory failure through glucolipotoxity. The relative importance of the transcription factors carbohydrate-responsive element binding protein (ChREBP), sterol-responsive element binding protein-1c (SREBP-1c), and upstream stimulatory factor (USF) in the induction of lipogenic genes by glucose remains unclear. By confocal imaging, we show that ChREBP translocates to the nucleus in MIN6 beta cells in response to glucose. Both ChREBP and SREBP-1c were required for the induction of the fatty acid synthase (FAS) promoter by glucose, and chromatin immunoprecipitation (ChIP) assay revealed that glucose induced the binding of both ChREBP and SREBP-1c to the FAS promoter without affecting USF2 binding. By contrast, ChIP assay revealed that high glucose prompted direct binding of ChREBP, but not SREBP-1c or USF2, to the liver-type pyruvate kinase (L-PK) promoter. This event was indispensable for the induction of the L-PK gene by glucose, as demonstrated by RNA silencing, single-cell promoter analysis, and quantitative real-time PCR. We conclude that ChREBP is a critical regulator of lipogenic genes in the beta cell and may play a role in the development of glucolipotoxicity and beta cell failure through alteration of gene expression in type 2 diabetes.

Active Transport, Cell Nucleus↗
PubMed1991

A. E. Braunstein Plenary Lecture. Nuclear skeleton, DNA domains and control of replication and transcription.

Chromosomal DNA is organized in loops or domains of about 100 kb. Their ends seem to be attached to special protein skeletal structures. The DNA-attachment sites can be subdivided into permanent and transient types. The permanent or constitutive attachment sites, which are retained in all types of cells (including those inactive in replication and transcription), either coincide with or are located close to replication origins. This observation provides a simple way for isolation of DNA fragments containing replication origins. Such fragments from the chicken alpha-globin gene domain and other regions of the chicken genome contain DNA sequences which interact with nuclear proteins present in dividing cells, but absent from non-dividing cells. Several new consensus sequences interacting with nuclear proteins were detected. The 5' end region of the alpha-globin gene domain containing a replication origin was found to possess enhancer activity lacking tissue specificity. Hence, the domain organization of DNA is related to the organization of replication process. Other sets of data indicate that the integrity of DNA domains is important for maintaining transcription within the domain. According to these data, even a single nick at an distance of about 100 kbp seems to be sufficient for blocking transcription within the whole domain at the stage of RNA elongation. Thus, topological integrity of DNA may be an important factor involved in formation of active chromatin.

Animals↗