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Autophagy is a highly conserved self-digestion process, which is essential for maintaining homeostasis and viability in response to nutrient starvation (1-4). Although the components of autophagy in the cytoplasm have been well studied (5,6), the molecular basis for the transcriptional and epigenetic regulation of autophagy is poorly understood. Here we identify co-activator-associated arginine methyltransferase 1 (CARM1) as a crucial component of autophagy in mammals. Notably, CARM1 stability is regulated by the SKP2-containing SCF (SKP1-cullin1-F-box protein) E3 ubiquitin ligase in the nucleus, but not in the cytoplasm, under nutrient-rich conditions. Furthermore, we show that nutrient starvation results in AMP-activated protein kinase (AMPK)-dependent phosphorylation of FOXO3a in the nucleus, which in turn transcriptionally represses SKP2. This repression leads to increased levels of CARM1 protein and subsequent increases in histone H3 Arg17 dimethylation. Genome-wide analyses reveal that CARM1 exerts transcriptional co-activator function on autophagy-related and lysosomal genes through transcription factor EB (TFEB). Our findings demonstrate that CARMl-dependent histone arginine methylation is a crucial nuclear event in autophagy, and identify a new signalling axis of AMPK-SKP2-CARM1 in the regulation of autophagy induction after nutrient starvation., To explore the importance of nuclear events in autophagy, we proposed that specific histone marks are involved in the epigenetic and transcriptional regulation of autophagy in the nucleus leading to [...]

Tumour oncogenes include transcription factors that co-opt the general transcriptional machinery to sustain the oncogenic state (1), but direct pharmacological inhibition of transcription factors has so far proven difficult (2). However, the transcriptional machinery contains various enzymatic cofactors that can be targeted for the development of new therapeutic candidates (3), including cyclin-dependent kinases (CDKs) (4). Here we present the discovery and characterization of a covalent CDK7 inhibitor, THZ1, which has the unprecedented ability to target a remote cysteine residue located outside of the canonical kinase domain, providing an unanticipated means of achieving selectivity for CDK7. Cancer cell-line profiling indicates that a subset of cancer cell lines, including human T-cell acute lymphoblastic leukaemia (T-ALL), have exceptional sensitivity to THZ1. Genome-wide analysis in Jurkat T-ALL cells shows that THZ1 disproportionally affects transcription of RUNX1 and suggests that sensitivity to THZ1 may be due to vulnerability conferred by the RUNX1 super-enhancer and the key role of RUNX1 in the core transcriptional regulatory circuitry of these tumour cells. Pharmacological modulation of CDK7 kinase activity may thus provide an approach to identify and treat tumour types that are dependent on transcription for maintenance of the oncogenic state., In an effort to discover new inhibitors of kinases that regulate gene transcription, we performed cell-based screening and kinase selectivity profiling of a library of known and novel ATP-site-directed kinase [...]

The cell-type-specific regulation of ribosomal RNA transcription is systematically studied. Several issues concerning the cell-type-specific requirement for rRNA production, the universality of Pol I transcription complex and the division of rDNA into regulatory subdomains are discussed.

Streptococcus pneumoniae normally resides in the human nasopharynx in a nondisease state. In response to unknown triggers this organism can descend to the lower respiratory tract and/or invade the bloodstream. Regulation and activation of virulence genes play essential roles in this process of disease development. Characterization of S. pneumoniae regulatory networks has been a recent area of interest, but despite inroads little is known about regulation of virulence genes in this pathogen. A putative transcriptional regulator in S. pneumoniae, mgrA, which exhibits homology to the virulence gene activator mga of group A streptococcus, was previously identified as a regulator that is required for development of pneumonia in a murine model. In this study we confirmed that mgrA plays a role in both nasopharyngeal carriage and pneumonia. Transcriptional profiling by microarray technology was used to show that mgrA acts as a repressor of the previously characterized rlrA pathogenicity islet. This is manifested phenotypically by a decrease in adherence to epithelial cells in tissue culture since the rlrA pathogenicity islet contains genes mediating adherence.

Regulation of transcription of the tryptophanase operon requires that translation of its leader peptide coding region, tnaC, be coupled with its transcription. We show in vitro that a transcription pause site exists at the end of the tnaC coding region and that translation of tnaC releases the paused transcription complex, coupling transcription with translation.

Results reveal that ToxT-dependent activation at tcpA promoter is mediated by the C-terminal domain of the RNA polymerase alpha subunit. ToxT interacts with Two adenine-rich regions of tcpA promoter assisted by the C-terminal domain of RNA polymerase alpha subunit.

Research reveals that the flagellar regulation in Pseudomonas aeruginosa follows two indepedent pathways mediated by an NtrC/NifA-type activator. In the typical pathway, transcription initiation occurs by upstream binding of activators and their interaction with RNA polymerase via looping in the fleSR promoter, whereas in the second the activator binds to polymerase without looping.

Paramutation is an interaction between alleles that leads to a heritable change in the expression of one allele. In B'/B-I plants, B-I (high transcription) always changes to B' (low transcription). The new B' allele retains the low expression state in the next generation and paramutates B-I at a frequency of 100%. Comparisons of the structure and expression of B' with that of a closely related allele that does not participate in paramutation demonstrated that transcription from the same promoter-proximal sequences is not sufficient for paramutation. Fine-structure recombination mapping localized sequences required for B' expression and paramutation. The entire 110 kb upstream of the B' transcription start site was cloned and sequenced and the recombination breakpoints were determined for 12 recombinant alleles. Sequences required for expression and paramutation mapped to distinct regions, 8.5-49 kb and 93-106 kb upstream of the B' transcription start site, respectively. Sequencing and DNA blot analyses indicate that the B' region required for paramutation is mostly unique or low copy in the maize genome. These results represent the first example of long-distance regulatory elements in plants and demonstrate that paramutation is mediated by long-distance cis and trans interactions.

Transcriptional activation of the inducible nitric oxide synthase (iNOS) gene requires multiple interactions of cis elements and trans-acting factors. Previous in vivo footprinting studies (Goldring CE, Reveneau S, Algarte M, and Jeannin JF. Nucleic Acids Res 24: 1682-1687, 1996) of the murine iNOS gene demonstrated lipopolysaccharide-inducible protection of guanines in the region -904/-883, which includes an E-box motif. In this report, by using site-directed mutagenesis of the -893/-888 E-box and correlating functional assays of the mutated iNOS promoter with upstream stimulatory factor (USF) DNA-binding activities, we demonstrate that the -893/-888 E-box motif is functionally required for iNOS regulation in murine mesangial cells and that USFs are in vivo components of the iNOS transcriptional response complex. Mutation of the E-box sequence augmented the iNOS response to interleukin-1[beta] (IL-1[beta]) in transiently transfected mesangial cells. Gel mobility shift assays demonstrated that USFs cannot bind to the -893/-888 E-box promoter region when the E-box is mutated. Cotransfection of USF-1 and USF-2 expression vectors with iNOS promoter-luciferase reporter constructs suppressed IL-1[beta]-simulated iNOS promoter activity. Cotransfection of dominant-negative USF-2 mutants lacking the DNA binding domain or cis-element decoys containing concatamers of the -904/-883 region augmented IL-1[beta] stimulation of iNOS promoter activity. Gel mobility shift assays showed that only USF-1 and USF-2 supershifted the USF protein-DNA complexes. These results demonstrated that USF binding to the E-box at -893/-888 serves to trans-repress basal expression and IL-1[beta] induction of the iNOS promoter. transcription; glomerulus; inducible nitric oxide synthase; promoter; inflammatory cytokines; transcription factors; upstream stimulatory factor

The regulation of transcription initiation of the promoter, by T7 RNA polymerase, occurs in multiple steps and is influenced by the promoter sequence and GTP concentration. The equilibrium constant of closed and open complexes, degree of promoter melting, the dissociation constant of initiating NTP, and rate of phosphodiester bond formation are regulated.

The transcription of the tir operon and the orf19 gene is mediated by Ler protein by overcoming the negative regulation of a repressor protein in Escherichia coli. Furthermore, cis-acting elements cesT and eae also regulate tir expression in response to environmental cues.

Results demonstrate the molecular basis for the exchange of coactivators for corepressors during development and homeostasis in murine cells is the discrimination by the nuclear receptor AF2 motif of the coactivator and corepressor interacting helices.

The biological effects induced by both protein kinase C-alpha and -delta were studied to determine whether they can be attributed to the cross-regulation between PKC-alpha and -delta. The findings demonstrated that the overexpression of PKC-alpha in Baf3 and 32D leads to an increase in protein levels and PKC-delta mRNA. The result was due to higher levels of mRNA stability and PKC-delta transcription.

Thioredoxin (TRX) is a pleiotropic cellular factor that has thiol-mediated redox activity and is important in regulation of cellular processes, including proliferation, apoptosis, and gene expression. The activity of several transcription factors is posttranslationally altered by redox modification(s) of specific cysteine residue(s). One such factor is nuclear factor (NF)-[Kappa]B, whose DNA-binding activity is markedly augmented by TRX treatment in vitro. Similarly, the DNA-binding activity of activator protein 1 (AP-1) is modified by a DNA repair enzyme, redox factor 1 (Ref-1), which is identical to a DNA repair enzyme, AP endonuclease. Ref-1 activity is in turn modulated by various redox-active compounds, including TRX. We here report the molecular cascade of redox regulation of AP-1 mediated by TRX and Ref-l. Phorbol 12-myristate 13 acetate efficiently translocated TRX into the HeLa cell nucleus where Ref-1 preexists. This process seems to be essential for AP-1 activation by redox modification because co-overexpression of TRX and Ref-1 in COS-7 cells potentiated AP-1 activity only after TRX was transported into the nucleus by phorbol 12-myristate 13 acetate treatment. To prove the direct active site-mediated association between TRX and Ref-1, we generated a series of substitution-mutant cysteine residues of TRX. In both an in vitro diamide-induced cross-linking study and an in vivo mammalian two-hybrid assay we proved that TRX can associate directly with Ref-1 in the nucleus; also, we demonstrated the requirement of cysteine residues in the TRX catalytic center for the potentiation of AP-1 activity. This report presents an example of a cascade in cellular redox regulation.

The molecular basis for the effect of hypoxia on animal life has gained the interest of biologists. Investigations have led to the recognition of hypoxia as a signal for energy conservation as well as an activator of genes responsible for the expression of the isoforms of glycolytic enzymes, glucose transporters that function better at low oxygen levels and erythropoietin (EPO). The hypoxia inducible factor 1 (HIF-1), on the other hand, has been identified as the transcription factor which specifically binds to the EPO enhancer. Results of further studies also proved HIF-1 as the global regulator of hypoxic gene expression.

The biochemical role of the upstream (A+T)-rich (ATR) regulatory region of the pRM and pR promoters of bacteriophage lambda is studied by utilizing templates with an inactivated pR promoter. Activation of the pRM promoter by the upstream regulatory region is not dependent on the (A+T) content of the region. Mutant pRM promoters with (G+C) substitutions exhibited normal biochemical activities indicating the sequence non-specific synthesis of nonoverlapping

The transcriptional regulation and organization of dnaK and groE operons of Chlamydia trachomatis was analyzed by the genetic sequencing and cloning of the genes from mouse pneumonitis (MoPn) biovars of the microbe. The dnaK operons of MoPn biovars of Chlamydias trachomatis is similar to the operons of Bacillus subtilis. The messenger ribonucleic acid of the operon also regulated in steady-state levels that is affected by heat shock. Nucleotide sequencing also indicated the presence of transcription sites in hrcA region of hrcA-grp-dnaK and the groES region of the groE operon.

The pathway from a zygote to an adult multicellular organism involves an immense number of components and interactions on each of several levels of suborganismal organization. There are thousands of [...]

A study of the Slp (sex-limited protein) gene in mouse liver indicates that the conserved regulatory element TTCCGGGC contains the methylation site at CpG (-121), the demethylation of this site occurring specifically in adult males. Expression of this gene requires transcription factors sensitive to methylation and DNA methylation. Methylation of the CpG site regulates binding between specific nuclear proteins and the conserved element.

FlgM gene transcription is negatively influenced by a class 2 flagellar mutation and a wild-type copy of the flgM gene. Residual expression of flgM in a delta-fliA strain results from read-through transcription from the adjoining flgA gene. These observations indicate that the FlgM gene is expressed from two promoters and may be classified as a member of both class 2 and class 3 flagellar gene.

An overproducing strain is used to purify the katF/rpoS gene derivative (sigma(super s)), which is a core controller of steady-phase gene expression in Escherichia coli. Monoclonal antibodies are generated by employing sigma(super s) as an immunogen. Homology to the sigma factor group is recognized by examining the order of the sigma(super s), whose property as a sigma factor is biochemically proved.

The human CD4 gene's relation to 5' transcription control sequences was explored and the transcription initiation region was identified. The CD4 core promoter does not have a classical 'TATA' or initiator positioning consensus sequence, but when coupled to the active simian virus 40 enhancer, it ensures accurate and effective transcription. By binding to an extraordinary proximal activator sequence and to a classical remote region, Ets transcription factors regulate CD4 gene expression.

Analysis of the obligate intracytoplasmic Rickettsia prowazeki indicates the presence of transcriptional regulation. The glt promoter in the P2 region of the microbe's deoxyribonucleic acid (DNA) indicated an inverted repeat sequence in the 47 bp region that is located upstream of the glt coding region. This DNA feature is a characteristic of microbes that possess positive transcriptional regulatory mechanisms.

A recombinase-based in vivo expression technology (IVET) was adapted to study the genes activated in Bacillus cereus ATCC 14579 during growth at low temperatures. The study demonstrated the feasibility of a promoter trap strategy for identifying cold-induced genes and also outlined the different processes involved in B. cereus cold acclimation.

The ferric uptake regulator (Fur)- and iron-regulated genes are identified in Listeria monocytogenes by DNA microanalysis using a fur mutant and its isogenic parent. The results have shown that Fur and PerR proteins have negatively regulated fur and that under iron-limited conditions, PeR is needed for the negative regulation of genes controlled by Fur.

An in vivo transposition system was used to identify novel regulators that influenced undecylprodigiosin (Red) production in Streptomyces coelicolor M145. The results from reverse transcription-PCR analysis revealed that RrdA negatively regulated Red production by controlling redD mRNA abundance, while no change was observed at the transcript level of the actinorhodin (Act)-specific activator gene, actII-orf4.

The organization of the RNA polymerase (RNAP) catalytic center, which has served as a robust, molecular engine that drives and controls the rate, processivity and fidelity of transcription is discussed.

Chromatin remodeling complexes (remodelers) have used the energy of ATP hydrolysis to move, destabilize, eject, or restructure nucleosomes. Many aspects of remodeler biology, their targeting, mechanism, regulation, shared and unique properties and specialization for particular biological processes are addressed.

Experiments are conducted to engineer complex riboswitches capable of sensing and responding to two small molecules according to Boolean logics AND and NAND. The results have highlighted the versatile capacity of the riboswitch to effect complex gene regulation carried out by modern protein transcription factors.

A time series of whole-genome transcription profiling of Escherichia coli K-12 W3110 is performed during a carbon-limited fed-batch process to identify dynamic sequence of diverse carbon limitation responses and provide global view of utilization of cellular and extracellular resources. The findings from the detailed study of the genetic regulation of bacterial metabolism could provide useful insight into manipulation of the carbon flux in technical production processes.

A novel network parameter that captures the high flow of information that passes through many miRNAs and TFs involved in composite feedback loops is discussed. A model is proposed in which feedback loops provide a mechanism for the highly coordinated and adapted control of gene batteries.