The multifunctionality of genome is suggested at some loci in different species but not well understood. Here we identified a DES-K16 region in an intron of the Kctd16 gene as the chromatin highly ...marked with epigenetic modifications of both enhancers (H3K4me1 and H3K27ac) and silencers (H3K27me3) in mouse spermatocytes. In vitro reporter gene assay demonstrated that DES-K16 exhibited significant enhancer activity in spermatocyte-derived GC-2spd(ts) and hepatic tumor-derived Hepa1-6 cells, and a deletion of this sequence in GC-2spd(ts) cells resulted in a decrease and increase of Yipf5 and Kctd16 expression, respectively. This was consistent with increased and decreased expression of Yipf5 and Kctd16, respectively, in primary spermatocytes during testis development. While known dual enhancer-silencers exert each activity in different tissues, our data suggest that DES-K16 functions as both enhancer and silencer in a single cell type, GC-2spd(ts) cells. This is the first report on a dual enhancer-silencer element which activates and suppresses gene expression in a single cell type.
•DES-K16 was marked with H3K4me1, H3K27ac, and H3K27me3 in mouse spermatocytes.•DES-K16 exhibited enhancer activity by in vitro reporter gene assay.•A deletion of DES-K16 down- and up-regulated Yipf5 and Kctd16 genes in GC-2spd(ts).•Yipf5 and Kctd16 expression was correlated to DES-K16 activity in the testis.•DES-K16 is a dual enhancer-silencer functioning in a single cell type.
Modification of nucleocytoplasmic proteins with O-GlcNAc regulates a wide variety of cellular processes and has been linked to human diseases. The enzymes O-GlcNAc transferase (OGT) and O-GlcNAcase ...(OGA) add and remove O-GlcNAc, but the mechanisms regulating their expression remain unclear. Here, we demonstrate that retention of the fourth intron of OGT is regulated in response to O-GlcNAc levels. We further define a conserved intronic splicing silencer (ISS) that is necessary for OGT intron retention. Deletion of the ISS in colon cancer cells leads to increases in OGT, but O-GlcNAc homeostasis is maintained by concomitant increases in OGA protein. However, the ISS-deleted cells are hypersensitive to OGA inhibition in culture and in soft agar. Moreover, growth of xenograft tumors from ISS-deleted cells is compromised in mice treated with an OGA inhibitor. Thus, ISS-mediated regulation of OGT intron retention is a key component in OGT expression and maintaining O-GlcNAc homeostasis.
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•O-GlcNAc transferase (OGT) expression is regulated by intron retention•OGT intron retention dynamically responds to O-GlcNAc levels•A conserved intronic splicing silencer (ISS) regulates OGT expression•The OGT-ISS is necessary for maintaining O-GlcNAc homeostasis
O-GlcNAc transferase (OGT) modifies cellular proteins, but the mechanisms that regulate OGT expression remain unclear. Park et al. show intron retention of the OGT transcript is responsive to cellular O-GlcNAc levels. They further define an intronic splicing silencer that is necessary to maintain O-GlcNAc homeostasis in cells and tumors.
Transposable elements, also known as transposons, are now recognized not only as parasitic DNA, the spread of which in the genome must be controlled by the host, but also as major players in genome ...evolution and regulation. Long interspersed element-1 (LINE-1, also known as L1), the only currently autonomous mobile transposon in humans, occupies 17% of the genome and generates inter- and intra-individual genetic variation, in some cases resulting in disease. However, how L1 activity is controlled and the function of L1s in host gene regulation are not completely understood. Here we use CRISPR-Cas9 screening strategies in two distinct human cell lines to provide a genome-wide survey of genes involved in the control of L1 retrotransposition. We identify functionally diverse genes that either promote or restrict L1 retrotransposition. These genes, which are often associated with human diseases, control the L1 life cycle at the transcriptional or the post-transcriptional level in a manner that can depend on the endogenous L1 nucleotide sequence, underscoring the complexity of L1 regulation. We further investigate the restriction of L1 by the protein MORC2 and by the human silencing hub (HUSH) complex subunits MPP8 and TASOR. HUSH and MORC2 can selectively bind evolutionarily young, full-length L1s located within transcriptionally permissive euchromatic environments, and promote deposition of histone H3 Lys9 trimethylation (H3K9me3) for transcriptional silencing. Notably, these silencing events often occur within introns of transcriptionally active genes, and lead to the downregulation of host gene expression in a HUSH-, MORC2-, and L1-dependent manner. Together, these results provide a rich resource for studies of L1 retrotransposition, elucidate a novel L1 restriction pathway and illustrate how epigenetic silencing of transposable elements rewires host gene expression programs.
Thousands of mutations are identified yearly. Although many directly affect protein expression, an increasing proportion of mutations is now believed to influence mRNA splicing. They mostly affect ...existing splice sites, but synonymous, non-synonymous or nonsense mutations can also create or disrupt splice sites or auxiliary cis-splicing sequences. To facilitate the analysis of the different mutations, we designed Human Splicing Finder (HSF), a tool to predict the effects of mutations on splicing signals or to identify splicing motifs in any human sequence. It contains all available matrices for auxiliary sequence prediction as well as new ones for binding sites of the 9G8 and Tra2-β Serine-Arginine proteins and the hnRNP A1 ribonucleoprotein. We also developed new Position Weight Matrices to assess the strength of 5′ and 3′ splice sites and branch points. We evaluated HSF efficiency using a set of 83 intronic and 35 exonic mutations known to result in splicing defects. We showed that the mutation effect was correctly predicted in almost all cases. HSF could thus represent a valuable resource for research, diagnostic and therapeutic (e.g. therapeutic exon skipping) purposes as well as for global studies, such as the GEN2PHEN European Project or the Human Variome Project.
The long-range interactions of cis-regulatory elements (cREs) play a central role in gene regulation. cREs can be characterized as accessible chromatin sequences. However, it remains technically ...challenging to comprehensively identify their spatial interactions. Here, we report a new method HiCAR (Hi-C on accessible regulatory DNA), which utilizes Tn5 transposase and chromatin proximity ligation, for the analysis of open-chromatin-anchored interactions with low-input cells. By applying HiCAR in human embryonic stem cells and lymphoblastoid cells, we demonstrate that HiCAR identifies high-resolution chromatin contacts with an efficiency comparable with that of in situ Hi-C over all distance ranges. Interestingly, we found that the “poised” gene promoters exhibit silencer-like function to repress the expression of distal genes via promoter-promoter interactions. Lastly, we applied HiCAR to 30,000 primary human muscle stem cells and demonstrated that HiCAR is capable of analyzing chromatin accessibility and looping using low-input primary cells and clinical samples.
•HiCAR captures chromatin accessibility and looping from the same input cells•HiCAR is cost-effective and can be applied to low-input clinical samples•HiCAR is compatible with SMART-seq for multi-omics analysis of DNA and RNA•The “poised” promoters exhibit silencer-like function via promoter-promoter looping
HiCAR utilizes Tn5 transposase and chromatin proximity ligation to capture open-chromatin-anchored interactions with low-input cells. It requires <10% sequencing depth of Hi-C to call high-resolution chromatin interactions. Interestingly, we found that the “poised” gene promoters exhibit silencer-like function to repress the expression of distal genes via promoter-promoter interactions.
•Relate analytically the transmission coefficient to the geometric parameters of the silencer permits the geometric optimization to achieve broadband high Transmission Loss (TL).•The staggered ...different array on a rectangular duct has the advantage of achieving flat transmission-loss curve over a considered frequency band.•By introducing more structural degrees of freedom in a fixed volume, it is possible to improve the broadband TL performance of the silencer.•The proposed design strategy has a potential to suppress broadband low frequency and can be easily generalized for waveguide with other cross-section shape, and thus can find broad applications in engineering projects.
In this work, we propose a general design strategy of passive acoustic treatments dealing with low frequency, broadband noise. The acoustic treatment is realized by arrays of Helmholtz resonators along the walls of the considered rectangular waveguide. The transfer matrix method is used to explicitly relate the geometric parameters of the resonators and acoustical response of the system. Based on this analytical model, an optimization is carried out to find the optimized structural design that minimizes the transmission coefficient in the considered frequency range. By using this design method, two specific designs are provided; their acoustic performances are validated by both simulations and measurements. The proposed design method can be easily generalized for waveguide with other cross-section shape, and thus can find broad applications in engineering projects which consider noise reductions in heat ventilation and air conditioning systems. Note that this work does not consider the effect of an air flow in the waveguide, however the design strategy still applies when the mean flow Mach number is sufficiently small.
•2D FEM to evaluate TL of perforated silencers with holes uniformly distributed.•A periodic square wave function was used to simulate the sound wave propagation.•The computational effort is much less ...than traditional FEM.•The developed formulation disregards the scattering phenomenon.•The developed formulation disregards the acoustic dissipation in the holes.
In this work, the two-dimensional finite element method approach is presented to evaluate sound transmission loss (TL) of perforated reactive silencers with holes uniformly distributed. In the hole domain, the sound pressure was described using a periodic positive square wave function to simplify the evaluation of sound waves in the azimuthal direction. Eight different models with perforated inner concentric tube are evaluated in terms of their TL. The proposed formulation results were verified using the traditional three-dimensional finite element method (FEM), showing good agreement. An additional verification was also performed using the three-dimensional FEM and the specific acoustic impedance of perforated inner concentric tube to couple the inner concentric duct with an expansion chamber. The periodic formulation and three-dimensional finite element analysis (FEA) used 4-node quadrilateral and 4-node tetrahedral elements, respectively. A final check was carried out with experimental analysis of 8 models and there was good agreement with results of the three numerical methods.
Genesis of novel gene regulatory modules is largely responsible for morphological and functional evolution. De novo generation of novel cis-regulatory elements (CREs) is much rarer than genomic ...events that alter existing CREs such as transposition, promoter switching or co-option. Only one case of de novo generation has been reported to date, in fish and without involvement of phenotype alteration. Yet, this event likely occurs in other animals and helps drive genetic/phenotypic variation.
Using a porcine model of spontaneous hearing loss not previously characterized we performed gene mapping and mutation screening to determine the genetic foundation of the phenotype. We identified a mutation in the non-regulatory region of the melanocyte-specific promoter of microphthalmia-associated transcription factor (MITF) gene that generated a novel silencer. The consequent elimination of expression of the MITF-M isoform led to early degeneration of the intermediate cells of the cochlear stria vascularis and profound hearing loss, as well as depigmentation, all of which resemble the typical phenotype of Waardenburg syndrome in humans. The mutation exclusively affected MITF-M and no other isoforms. The essential function of Mitf-m in hearing development was further validated using a knock-out mouse model.
Elimination of the MITF-M isoform alone is sufficient to cause deafness and depigmentation. To our knowledge, this study provides the first evidence of a de novo CRE in mammals that produces a systemic functional effect.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The ETS transcription factor PU.1 plays an essential role in blood cell development. Its precise expression pattern is governed by cis-regulatory elements (CRE) acting at the chromatin level. CREs ...mediate the fine-tuning of graded levels of PU.1, deviations of which can cause acute myeloid leukemia. In this review, we perform an in-depth analysis of the regulation of PU.1 expression in normal and malignant hematopoiesis. We elaborate on the role of trans-acting factors and the biomolecular interplays in mediating local chromatin dynamics. Moreover, we discuss the current understanding of CRE bifunctionality exhibiting enhancer or silencer activities in different blood cell lineages and future directions toward gene-specific chromatin-targeted therapeutic development.
•A natural ventilation window that achieves global passive control of transformer noise.•A coupled coiled-up structure that reduces noise at 4 frequencies simultaneously.•Verification of the ...effectiveness of the natural ventilation window with a scale model.
For transformers located inside rooms, openings in the walls are often required for ventilation and heat dissipation with the result that transformer noise radiates to the outside. A soundproof window for indoor transformers is proposed in this paper, which provides both air circulation and noise reduction simultaneously. A silencer consisting of two coupled tubes with different cross sections is designed and coiled up in space to minimize the thickness of the structure. With carefully chosen parameters, just one such silencer can achieve sound attenuation at up to 4 frequencies. With a combination of a staggered window and specially designed silencers, effective noise reduction is obtained at 100 Hz, 200 Hz, 300 Hz, 400 Hz and 500 Hz, where harmonic components contribute the most to the transformer noise. The experimental results with a 1:4 scale down model show the feasibility of the proposed design.
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