Hierarchical and stage-specific regulation of murine cardiomyocyte maturation by serum response factor

After birth, cardiomyocytes (CM) acquire numerous adaptations in order to efficiently pump blood throughout an animal’s lifespan. How this maturation process is regulated and coordinated is poorly understood. Here, we perform a CRISPR/Cas9 screen in mice and identify serum response factor (SRF) as a key regulator of CM maturation. Mosaic SRF depletion in neonatal CMs disrupts many aspects of their maturation, including sarcomere expansion, mitochondrial biogenesis, transverse-tubule formation, and cellular hypertrophy. Maintenance of maturity in adult CMs is less dependent on SRF. This stage-specific activity is associated with developmentally regulated SRF chromatin occupancy and transcriptional regulation. SRF directly activates genes that regulate sarcomere assembly and mitochondrial dynamics. Perturbation of sarcomere assembly but not mitochondrial dynamics recapitulates SRF knockout phenotypes. SRF overexpression also perturbs CM maturation. Together, these data indicate that carefully balanced SRF activity is essential to promote CM maturation through a hierarchy of cellular processes orchestrated by sarcomere assembly

Hierarchical and stage-specific regulation of murine cardiomyocyte maturation by serum response factor

After birth, cardiomyocytes (CM) acquire numerous adaptations in order to efficiently pump blood throughout an animal’s lifespan. How this maturation process is regulated and coordinated is poorly understood. Here, we perform a CRISPR/Cas9 screen in mice and identify serum response factor (SRF) as a key regulator of CM maturation. Mosaic SRF depletion in neonatal CMs disrupts many aspects of their maturation, including sarcomere expansion, mitochondrial biogenesis, transverse-tubule formation, and cellular hypertrophy. Maintenance of maturity in adult CMs is less dependent on SRF. This stage-specific activity is associated with developmentally regulated SRF chromatin occupancy and transcriptional regulation. SRF directly activates genes that regulate sarcomere assembly and mitochondrial dynamics. Perturbation of sarcomere assembly but not mitochondrial dynamics recapitulates SRF knockout phenotypes. SRF overexpression also perturbs CM maturation. Together, these data indicate that carefully balanced SRF activity is essential to promote CM maturation through a hierarchy of cellular processes orchestrated by sarcomere assembly

Study on Temperature Characteristics of Lubrication Film of Valve Plate Pair in Axial Piston Pumps

The film temperature distribution of the valve plate pair in axial piston pumps affects its lubrication, leakage, and friction. In order to investigate the film temperature distribution of the valve plate pair in axial piston pumps, a test platform was constructed including three displacement sensors for the oil film thickness and eleven thermocouples for the film temperature distribution of the valve plate pair. An accurate film shape model of the valve plate pair was built according to the three-point film thickness test data. Based on the film shape model, the film temperature model of the valve plate pair was developed considering the viscous oil temperature characteristics, the energy loss caused by leakage and viscous friction in the film, and the heat conduction among the oil, cylinder block, and valve plate. The influence of different swash plate tilt angles and operating pressures on the valve plate film temperature was studied. The test results indicate that the film temperature of the valve plate pair increases as the working pressure and swash plate tilt angle increase. The theoretical and experimental absolute errors of the film temperature in the circumferential range [−60°, 60°] of the valve plate high-pressure side are less than 3.5 °C. As the swash plate tilt angle varies from 12° to 16° and working pressure from 3 MPa to 7 MPa, the minimum film thickness position and the maximum temperature point move accordingly in the circumferential range [−15°, 5°] of the valve plate pair

Acupuncture for functional dyspepsia: Bayesian meta-analysis

Background: Acupuncture stands out as a prominent complementary and alternative medicine therapy employed for functional dyspepsia (FD). We conducted a Bayesian meta-analysis to ascertain both the relative effectiveness and safety of various acupuncture methods in the treatment of functional dyspepsia. Methods: We systematically searched eight electronic databases, spanning from their inception to April 2023. The eligibility criteria included randomized controlled trials investigating acupuncture treatments for FD. Study appraisal was conducted using the Cochrane risk of bias tool. Pairwise and network meta-analyses were conducted using RevMan 5.3 and ADDIS V.1.16.6 software. Bayesian network meta-analysis was performed to compare and rank the efficacy of different acupuncture therapies for FD symptoms. Results: This study found that combining different acupuncture methods or using acupuncture in conjunction with Western medicine is more effective in improving symptoms of functional dyspepsia compared to using Western medicine alone. According to the comprehensive analysis results, notably, the combination of Western medicine and acupuncture exhibited superior efficacy in alleviating early satiation and postprandial fullness symptoms. For ameliorating epigastric pain, acupuncture combined with moxibustion proved to be the most effective treatment, while moxibustion emerged as the optimal choice for addressing burning sensations. Warming needle was identified as the preferred method for promoting motilin levels. Conclusion: The findings of this study demonstrate that acupuncture, both independently and in conjunction with other modalities, emerged as a secure and effective treatment option for patients with functional dyspepsia

Mapping cell type-specific transcriptional enhancers using high affinity, lineage-specific Ep300 bioChIP-seq

Understanding the mechanisms that regulate cell type-specific transcriptional programs requires developing a lexicon of their genomic regulatory elements. We developed a lineage-selective method to map transcriptional enhancers, regulatory genomic regions that activate transcription, in mice. Since most tissue-specific enhancers are bound by the transcriptional co-activator Ep300, we used Cre-directed, lineage-specific Ep300 biotinylation and pulldown on immobilized streptavidin followed by next generation sequencing of co-precipitated DNA to identify lineage-specific enhancers. By driving this system with lineage-specific Cre transgenes, we mapped enhancers active in embryonic endothelial cells/blood or skeletal muscle. Analysis of these enhancers identified new transcription factor heterodimer motifs that likely regulate transcription in these lineages. Furthermore, we identified candidate enhancers that regulate adult heart- or lung- specific endothelial cell specialization. Our strategy for tissue-specific protein biotinylation opens new avenues for studying lineage-specific protein-DNA and protein-protein interactions. DOI: http://dx.doi.org/10.7554/eLife.22039.00

Interrogating translational efficiency and lineage-specific transcriptomes using ribosome affinity purification

Transcriptional profiling is a useful strategy to study development and disease. Approaches to isolate RNA from specific cell types, or from specific cellular compartments, would extend the power of this strategy. Previous work has shown that isolation of genetically tagged ribosomes (translating ribosome affinity purification; TRAP) is an effective means to isolate ribosome-bound RNA selectively from transgene-expressing cells. However, widespread application of this technology has been limited by available transgenic mouse lines. Here we characterize a TRAP allele (Rosa26[superscript fsTRAP]) that makes this approach more widely accessible. We show that endothelium-specific activation of Rosa26[superscript fsTRAP] identifies endothelial cell-enriched transcripts, and that cardiomyocyte-restricted TRAP is a useful means to identify genes that are differentially expressed in cardiomyocytes in a disease model. Furthermore, we show that TRAP is an effective means for studying translational regulation, and that several nuclear-encoded mitochondrial genes are under strong translational control. Our analysis of ribosome-bound transcripts also shows that a subset of long intergenic noncoding RNAs are weakly ribosome-bound, but that the majority of noncoding RNAs, including most long intergenic noncoding RNAs, are ribosome-bound to the same extent as coding transcripts. Together, these data show that the TRAP strategy and the Rosa26[superscript fsTRAP] allele will be useful tools to probe cell type-specific transcriptomes, study translational regulation, and probe ribosome binding of noncoding RNAs.American Heart AssociationNational Heart, Lung, and Blood Institute (U01HL098166)National Heart, Lung, and Blood Institute (U01HL098188)National Heart, Lung, and Blood Institute (R01HL095712

VEGF amplifies transcription through ETS1 acetylation to enable angiogenesis

Release of promoter-proximally paused RNA polymerase II (RNAPII) is a recently recognized transcriptional regulatory checkpoint. The biological roles of RNAPII pause release and the mechanisms by which extracellular signals control it are incompletely understood. Here we show that VEGF stimulates RNAPII pause release by stimulating acetylation of ETS1, a master endothelial cell transcriptional regulator. In endothelial cells, ETS1 binds transcribed gene promoters and stimulates their expression by broadly increasing RNAPII pause release. 34 VEGF enhances ETS1 chromatin occupancy and increases ETS1 acetylation, enhancing its binding to BRD4, which recruits the pause release machinery and increases RNAPII pause release. Endothelial cell angiogenic responses in vitro and in vivo require ETS1-mediated transduction of VEGF signaling to release paused RNAPII. Our results define an angiogenic pathway in which VEGF enhances ETS1–BRD4 interaction to broadly promote RNAPII pause release and drive angiogenesis

Dynamic changes in P300 enhancers and enhancer-promoter contacts control mouse cardiomyocyte maturation

Cardiomyocyte differentiation continues throughout murine gestation and into the postnatal period, driven by temporally regulated expression changes in the transcriptome. The mechanisms that regulate these developmental changes remain incompletely defined. Here, we used cardiomyocyte-specific ChIP-seq of the activate enhancer marker P300 to identify 54,920 cardiomyocyte enhancers at seven stages of murine heart development. These data were matched to cardiomyocyte gene expression profiles at the same stages and to Hi-C and H3K27ac HiChIP chromatin conformation data at fetal, neonatal, and adult stages. Regions with dynamic P300 occupancy exhibited developmentally regulated enhancer activity, as measured by massively parallel reporter assays in cardiomyocytes in vivo, and identified key transcription factor-binding motifs. These dynamic enhancers interacted with temporal changes of the 3D genome architecture to specify developmentally regulated cardiomyocyte gene expressions. Our work provides a 3D genome-mediated enhancer activity landscape of murine cardiomyocyte development