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Results for “Transcription Initiation, Genetic”

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At least 19 recordsLinked to original sources
PubMed2025

BRD2 bridges TFIID and MOF-H4K16ac-containing nucleosomes to promote transcriptional initiation.

Members of the bromodomain and extraterminal domain (BET) protein family play a central role in transcription by RNA polymerase II (RNA Pol II). Small-molecule inhibitors that block interaction between BET bromodomains and acetylated histones have been developed for disease therapeutics. However, the BET protein BRD4 does not require bromodomains to perform its major transcriptional elongation control, and mechanisms by which other BET proteins regulate RNA Pol II remain insufficiently understood. Addressing the disparity between pan-BET degraders and BRD4-specific depletion, we report that the BET protein BRD2 generally functions to promote transcriptional initiation in a bromodomain-dependent manner at both promoters and enhancers in human cell lines. We demonstrate that BRD2 bromodomains preferentially bind to histone H4 harboring MOF-mediated H4K16ac, while the BRD2 C-terminal domain facilitates recruitment of TFIID. Our studies provide mechanistic insight into distinct roles for BRD2 and BRD4 in transcriptional initiation and elongation control for proper regulation of gene expression.

Humans↗
PubMed2024

The HIV-1 Transcriptional Program: From Initiation to Elongation Control.

A large body of work in the last four decades has revealed the key pillars of HIV-1 transcription control at the initiation and elongation steps. Here, I provide a recount of this collective knowledge starting with the genomic elements (DNA and nascent TAR RNA stem-loop) and transcription factors (cellular and the viral transactivator Tat), and later transitioning to the assembly and regulation of transcription initiation and elongation complexes, and the role of chromatin structure. Compelling evidence support a core HIV-1 transcriptional program regulated by the sequential and concerted action of cellular transcription factors and Tat to promote initiation and sustain elongation, highlighting the efficiency of a small virus to take over its host to produce the high levels of transcription required for viral replication. I summarize new advances including the use of CRISPR-Cas9, genetic tools for acute factor depletion, and imaging to study transcriptional dynamics, bursting and the progression through the multiple phases of the transcriptional cycle. Finally, I describe current challenges to future major advances and discuss areas that deserve more attention to both bolster our basic knowledge of the core HIV-1 transcriptional program and open up new therapeutic opportunities.

HIV-1↗
PubMed2026

Single-nucleotide transcription start sites profiling via Nascent Strand-Specific RNA sequencing uncovers IFN-γ-induced promoter dynamics.

Transcriptional regulation is a highly dynamic process in which nascent RNAs provide the most immediate readout of transcriptional activity. Precise mapping of transcription start sites (TSSs) is therefore critical for understanding promoter architecture and gene regulation, yet remains technically challenging. Here, we introduce Nascent Strand-Specific RNA sequencing (NSS-seq), a robust and streamlined method for genome-wide profiling of the capped 5' ends of nascent RNAs. By directly capturing transcription initiation events, NSS-seq overcomes the temporal delay inherent to conventional RNA-seq and enables time-resolved interrogation of transcriptional dynamics. Applied to interferon-γ (IFN-γ)-stimulation, NSS-seq uncovers previously unrecognized IFN-γ-responsive genes and transient transcription factor activation patterns underlying interferon-mediated tumor-suppressive functions. Together, NSS-seq provides a cost-effective and technically accessible platform for dissecting promoter-level regulatory dynamics during cellular responses.

Promoter Regions, Genetic↗
PubMed2026

Alternative tandem transcription initiation links noncoding variants to human disease through translational control.

Alternative tandem transcription initiation is a pervasive mechanism of gene regulation, yet its genetic impact on human disease remains largely unknown. Here, we systematically quantify the genetic regulation of alternative tandem transcription initiation across 25,859 samples from 49 normal human tissues and 33 tumor tissues. We identify approximately 0.4 million genetic variants associated with alternative transcription initiation in 5295 genes, with 32% operating independently of gene expression. Moreover, we discover 2238 multi-tissue alternative tandem transcription initiation outliers enriched for rare deleterious promoter and 5' UTR variants, demonstrating that both common and rare variants modulate transcription initiation. Strikingly, 74% of disease variants that colocalize with genetic variants regulating alternative transcription initiation cannot be identified through expression quantitative trait loci. Transcriptome-wide association studies identify 614 disease susceptibility genes associated with alternative transcription initiation, including known cancer drivers such as MAFF and MLLT10. Functional validation uncovers OSGEP as a breast cancer risk gene, where the alternative allele lengthens the 5' UTR and reduces protein abundance through upstream open reading frame-mediated translation repression, and suppresses breast cancer cell proliferation. Our findings establish alternative transcription initiation as a major, underappreciated mechanism associating noncoding variation with disease, providing a critical resource for interpreting disease risk loci.

Humans↗
PubMed2026

Oriented binding of transcription factors to nucleosomes remodels chromatin at human promoters.

Transcription factors (TFs) can access nucleosomes via five distinct modes: gyre-spanning, periodic-binding, dyad-binding, and end-binding modes as well as an oriented binding mode, where the TF binding motif shows orientational preference relative to the nucleosome. Here, we report the first structure of an oriented TF:nucleosome complex, where two ELF2 proteins bind to a double motif located at superhelical location +4, unwinding four helical turns of DNA from the nucleosome. We further show that unlike previously described pioneer factors, ELF2 is able to occupy all of its unmethylated, high-affinity double motifs in vivo. Motifs of ELF2 and another oriented nucleosome binder, YY1, are highly enriched downstream of transcription start sites (TSSs) of highly expressed genes, with the motifs oriented in such a way that the TSS becomes accessible upon TF binding. Our results suggest that oriented binding may be generally important for high transcriptional activity.

Nucleosomes↗
PubMed2026

Activation of mTOR pathway by human cytomegalovirus promoting host ribosomal protein expression by coordinated transcriptional and translational controls.

Human cytomegalovirus (HCMV) profoundly reprograms host transcription and RNA metabolism, yet its impact on transcription start site (TSS) regulation of host genes remains poorly understood. Here, we employed NanoCap Analysis of Gene Expression sequencing (NanoCAGE-seq) to investigate HCMV-driven changes in alternative TSS usage across the host transcriptome. We identified widespread TSS switching, with ribosomal protein genes (RPGs) emerging as a highly enriched category. Alternative TSS usage produced isoforms with distinct 5'untranslated regions (UTRs), thereby altering cis-regulatory elements that shape translational efficiency. Integrative transcriptomic and proteomic analyses revealed a paradoxical accumulation of RPG proteins despite transcriptional downregulation during infection. Using 5' Rapid Amplification of cDNA Ends (5'RACE), we characterized four RPGs of RPL4, RPS11, RPS23, and RPS24 that generated 5'UTR variants through alternative TSS usage. Notably, isoforms containing a 5'terminal oligopyrimidine (5'TOP) motif were significantly enriched, correlating with mTOR activation induced by HCMV. Functional assays with bicistronic reporter constructs in HEK293 cells and infection models in human embryonic lung fibroblasts demonstrated that the RPL4 5'TOP isoform exhibited enhanced mTORC1-driven translation compared with non-5'TOP counterparts. Importantly, RPL4 upregulation facilitated viral protein synthesis and boosted production of infectious virions. Together, our findings reveal that dynamic TSS switching of RPGs provides a simple, yet effective, mechanism for fine-tuning mTORC1-responsive translation. By co-opting host transcriptional and translational programs, HCMV enhances ribosome function to optimize the cellular environment for productive viral replication.

Humans↗
PubMed2026

The circadian clock proteins PRR modulate root hair development via the RHD6/RSL module in Arabidopsis.

Root hairs, derived from trichoblasts, are critical for plant growth and environmental adaptation. Although environmental cues are known to influence root hair development, how endogenous timing systems such as the circadian clock integrate into the core transcriptional network governing root hair formation remains unclear. Here, we show that the circadian clock-associated protein PSEUDO-RESPONSE REGULATOR5 (PRR5) physically interacts with ROOT HAIR DEFECTIVE6 (RHD6) and RHD6 LIKE1 (RSL1), two basic helix-loop-helix transcription factors essential for root hair initiation. Genetic analyses suggest that PRR proteins contribute to root hair development under long-day conditions in Arabidopsis thaliana. Simultaneous disruption of PRR5, PRR7, and PRR9 results in defective root hairs, whereas PRR5 overexpression markedly increases root hair density and length. Transcriptomic and RT-qPCR analyses reveal that PRRs enhance the expression of RHD6, RSL1, and multiple downstream root hair-responsive genes, while modulating their temporal expression patterns. Furthermore, PRR5-mediated root hair promotion requires RHD6/RSL1, and PRR proteins enhance RHD6-dependent activation of the RSL4 promoter. PRRs also contribute to root hair development under phosphate-deficient and salt-stress conditions. Together, these findings establish a molecular framework in which PRR proteins regulate the RHD6/RSL network to coordinate root hair development and environmental responses.

Arabidopsis↗
PubMed1978

Gene expression in Escherichia coli B/r during partial rifampicin-mediated restrictions of transcription initiation.

The antibiotic rifampicin inhibits transcription initiation, but not the elongation and completion of nascent RNA transcripts. Addition of low concentrations of rifampicin only partially blocks initiation but at the same time specifically alters the general pattern of transcription in the culture. The transcription of genes specifying the beta and beta' subunits of RNA polymerase, and to a lesser extent of the genes specifying the RNA and protein components of the ribosome, was specifically stimulated relative to total transcription. In contrast, the transcription of the lactose operon was selectively reduced. These results are consistent with the ideas that the level of expression of the genes specifying the beta and beta' subunits is sensitive to the general rate of RNA synthesis in the culture, and that the expression of the beta and beta' RNA polymerase genes is related to the expression of ribosome component genes.

DNA-Directed RNA Polymerases↗
PubMed1978

Studies on the regulation of RNA synthesis in neuronal and glial nuclei isolated from rat brain.

In searching for regulatory mechanisms involved in the cell-specific neuronal and glial transcription a cell-free transcriptional system has been developed using neuronal and glial rat brain chromatin and partially purified neuronal and glial nuclear rat brain RNA polymerases. Both free and chromatin-bound (engaged) neuronal and glial RNA polymerase fractions were separated from isolated neuronal and glial rat brain nuclei to determine their transcriptive efficiency. A double number of RNA initiation sites was measured on the neuronal when compared to the glial chromatin, independently of whether the neuronal or the glial RNA polymerase preparation was used for the determination. Structural modification of the neuronal and glial chromatin template by acetylation with acetyl-coenzyme A leads to an increase of the total number of RNA initiation sites available for exogenously added rat brain RNA polymerase. This indicates that acetylation of chromatin-bound proteins is capable to render primarily restricted gene sequences transcriptable. A positive correlation exists between the extent of acetate uptake by neuronal and glial chromatin-bound histone fractions and the extent of the increase of the number of RNA initiation sites is specifically related to histone acetylation rather than to acetylation of any other chromatin protein. Significant information in this respect could be achieved by dissociation of chromatin into its principal components and selectively reconstituting DNA with specifically acetylated histone and non-histone proteins.

Animals↗
PubMed1977

Transcription of insertion elements IS1 and IS2 in vitro.

Insertion elements IS1 and IS2 integrated within the gal operator-promoter region, an IS1 element in gene galT and insertions IS1 and IS2 integrated in the xycIIOP region of phage lambda were transcribed in vitro with E. coli RNA-polymerase. The insertion elements are transcribed exclusively by polymerase molecules started at the gal promoter and the lambdaPR promoter respectively. No promoter exists on IS1 or IS2 which can be recognized by RNA-polymerase in the pure in vitro transcription system used. Both insertions apparently are transcribed with a lower elongation rate than gal operon DNA or lambdaDNA. RNAs transcribed from the termini of IS1 and IS2 respectively were analysed by hybridization experiments. They are different in sequence.

Base Sequence↗
PubMed1978

Nucleotide sequence of cro, cII and part of the O gene in phage lambda DNA.

A nucleotide sequence comprising 960 base pairs of bacteriophage lambda DNA has been determined. The sequence includes the entire genes of the regulatory proteins cro and cII, and part of the O gene, together with control elements for their transcription and translation. The right-hand boundaries of the lambdaimm434 and lambdaimm21 substitutions and the cy42 mutation have been located.

Base Sequence↗
PubMed2026

Transcription Start Regions in PTU-intergenic regions drive cell cycle-dependent transcriptional activation events in Leishmania donovani.

Leishmania displays an unconventional mode of transcription, with long clusters of genes being transcribed polycistronically from Transcription Start Regions (TSRs), being processed into monocistronic units prior to translation. It has long been believed that transcription is constitutive: failure to identify consensus sequences across TSRs (except a GT-rich motif supporting transcription in Trypanosoma brucei) and absence of canonical eukaryotic transcription factors led to the conclusion that regulation is primarily post-transcriptional, with epigenetics playing a role in triggering transcription initiation. This study stems from our previous findings identifying a few genes to be activated in a cell cycle-dependent manner. Using nuclear run-on assays to analyze nascent transcripts of two chromosomes, chromosomes 2 and 14, we find that while most genes are constitutively transcribed, a subset of genes gets activated at specific cell cycle stages. Reporter assays reveal that this transcriptional activation is driven by the regions immediately upstream of the genes. Sequence analyses of these TSRs lying in polycistronic intergenic regions (PIRs) uncovered a 10-mer GT-rich motif, in synchrony with earlier findings in T. brucei identifying a GT-rich motif at bidirectional TSRs. We also identify a second 25-mer motif at these TSRs, and deletion analyses find this motif to be critical for regulating gene expression. The findings of this study reveal that transcriptional events in these unicellular parasites are more complex than believed thus far: not all transcriptional events are constitutive, polycistronic transcription is not the only mode of transcription, and cis-acting sequence elements regulate at least some transcriptional events in these parasites.IMPORTANCEEndemic to 90 countries, Leishmania parasites cause a spectrum of diseases called Leishmaniases. No vaccines for human use are available to date, and the drugs currently used to treat the disease are expensive, have toxic side effects, and have complex administration regimens, with emerging drug resistance compounding problems. Researchers continue to investigate Leishmania cellular processes, with the hope of uncovering new therapeutic target sites. Gene regulation in these parasites is unusual, being modulated by various mechanisms, including epigenetic modifications, gene dosage, and post-transcriptional processing. Transcription is typically polycistronic and constitutive, initiating from Transcription Start Regions (TSRs) lying upstream of the first gene in the polycistronic transcription unit (PTU). The work presented here reveals that a subset of genes is transcribed monocistronically in a cell cycle-dependent manner from Transcription Start Regions lying in the PTU-intergenic regions (PIRs), underscoring the complexities of gene regulation in these parasites.

Leishmania donovani↗
PubMed1979

DNA-directed in vitro synthesis of beta-galactosidase. Purification and characterization of stimulatory factors in an ascites extract.

Previous studies have described a partially defined system for the DNA-directed in vitro synthesis of beta-galactosidase (Kung, H.F., Redfield, B., Treadwell, B.V., Eskin, B., Spears, C., and Weissbach, H. (1977) J. Biol. Chem. 252, 6889-6894). An Ehrlich ascites extract was shown in these in vitro studies to acylate Escherichia coli tRNA with 13 amino acids, and the ascites extract was used in place of the corresponding 13 E. coli aminoacyl-tRNA synthetases. The present studies indicate that the ascites extract is supplying an additional protein factor, besides the aminoacyl-tRNA synthetases, that stimulates the DNA-directed synthesis of beta-galactosidase. The protein factor has been highly purified and may be functioning by protecting mRNA against degradation. In addition, NAD or T4 DNA ligase stimulates the synthesis of beta-galactosidase in the partially defined system.

Amino Acyl-tRNA Synthetases↗
PubMedDate not supplied

[Initiation in a polyribosome-dependent protein-synthesizing cell-free system from Saccharomyces (author's transl)].

A method to prepare polyribosomes from yeasts by using the french-press is described. The highest yield of polyribosomes was derived from late log-phase cells. These polyribosomes, incubated in a cell-free system, were able to reinitiate protein synthesis, which was shown by inhibiting aminoacid incorporation by aurintricarboxylic acid, edeine and sodiumfluoride. We developed the translational system in order to look for the optimal ion-conditions of a DNA-dependent protein-synthesizing system. We found out that at the optimal MgCl2-concentration (6 mM) protein synthesis was strongly inhibited by Mangan ions which are required for transcription in yeast. If protein-synthesis was carried out with 2 mM and 3 mM MgCl2 maximal aminoacid incorporation was observed at 2 mM and 1.5 mM MnCl2.

Aurintricarboxylic Acid↗
PubMed1979

Characterization of the 5'-terminal structure of simian virus 40 early mRNA's.

RPC-5 reverse-phase chromatography has been used to isolate fragments of simian virus 40 DNA generated by appropriate digestions with restriction endonucleases. Ten specific DNA fragments, mapping successively in a counterclock-wise direction from 0.67 to 0.515 on the simian virus 40 genome, were each hybridized to cytoplasmic mRNA obtained during the early phase of simian virus 40 infection. Primer extension methods with reverse transcriptase were used to characterize the 5' ends of two species of viral mRNA which were fractionated on sucrose gradients. Analysis of the complementary DNA products demonstrated the presence of two different spliced structures of simian virus 40 early mRNA's, both of which had the same 5'-end sequences (AUU), located at residues 18 to 20 on the viral genome. The mRNA for small-t contained a segment 588 bases in length (residues 18 to 605) spliced to residues 672. A 66-nucleotide segment rich in adenine-thymine was spliced out of this mRNA. The mRNA for large-T contained a segment 308 bases in length (residues 18 to 325) which is also spliced to residue 672. A 346-base segment was spliced from this mRNA. The results suggest that there are two levels for control of genetic expression. One would be the regulation of initiation of transcription at a common promoter; the other involves post-transcriptional splicing.

Base Sequence↗
PubMed1977

Thermal denaturation and template activities of reconstituted DNA-histone complexes.

Reconstituted complexes of DNA with histone were prepared by salt-and-urea step gradient dialysis. The DNA was complexed with histone H1, with the combination of the other four histones H2A, H2B, H3 and H4, and with whole histones. These DNA-histone complexes were purified by Bio-Gel column chromatography, and the weight ratio of histone-to-DNA was determined in each complex. The thermal denaturation profile and nuclease digestion pattern of DNA-histone H2A, H2B, H3 and H4 complex were compatible with those of the polynucleosome structure of chromatin. The template activities for transcription were compared in these DNA-histone complexes by separately measuring initiation reaction and chain elongation. The binding of histone H1 to DNA strongly inhibited the initiation, while the binding of the combination of the other four histones to DNA partially inhibited the initiation and chain elongation. The binding characteristics are discussed with regard to the role of histone H1 and the other four histones in chromatin structure and template activity.

Animals↗