Chromatin profiling reveals TFAP4 as a critical transcriptional regulator of bovine satellite cell differentiation.

BACKGROUND: Satellite cells are myogenic precursor cells in adult skeletal muscle and play a crucial role in skeletal muscle regeneration, maintenance, and growth. Like embryonic myoblasts, satellite cells have the ability to proliferate, differentiate, and fuse to form multinucleated myofibers. In this study, we aimed to identify additional transcription factors that control gene expression during bovine satellite cell proliferation and differentiation. RESULTS: Using chromatin immunoprecipitation followed by sequencing, we identified 56,973 and 54,470 genomic regions marked with both the histone modifications H3K4me1 and H3K27ac, which were considered active enhancers, and 50,956 and 59,174 genomic regions marked with H3K27me3, which were considered repressed enhancers, in proliferating and differentiating bovine satellite cells, respectively. In addition, we identified 1,216 and 1,171 super-enhancers in proliferating and differentiating bovine satellite cells, respectively. Analyzing these enhancers showed that in proliferating bovine satellite cells, active enhancers were associated with genes stimulating cell proliferation or inhibiting myoblast differentiation whereas repressed enhancers were associated with genes essential for myoblast differentiation, and that in differentiating satellite cells, active enhancers were associated with genes essential for myoblast differentiation or muscle contraction whereas repressed enhancers were associated with genes stimulating cell proliferation or inhibiting myoblast differentiation. Active enhancers in proliferating bovine satellite cells were enriched with binding sites for many transcription factors such as MYF5 and the AP-1 family transcription factors; active enhancers in differentiating bovine satellite cells were enriched with binding sites for many transcription factors such as MYOG and TFAP4; and repressed enhancers in both proliferating and differentiating bovine satellite cells were enriched with binding sites for NF-kB, ZEB-1, and several other transcription factors. The role of TFAP4 in satellite cell or myoblast differentiation was previously unknown, and through gene knockdown and overexpression, we experimentally validated a critical role for TFAP4 in the differentiation and fusion of bovine satellite cells into myofibers. CONCLUSIONS: Satellite cell proliferation and differentiation are controlled by many transcription factors such as AP-1, TFAP4, NF-kB, and ZEB-1 whose roles in these processes were previously unknown in addition to those transcription factors such as MYF5 and MYOG whose roles in these processes are widely known.

Molecular and Cellular Mechanisms of Intramuscular Fat Development and Growth in Cattle.

Intramuscular fat, also referred to as marbling fat, is the white fat deposited within skeletal muscle tissue. The content of intramuscular fat in the skeletal muscle, particularly the longissimus dorsi muscle, of cattle is a critical determinant of beef quality and value. In this review, we summarize the process of intramuscular fat development and growth, the factors that affect this process, and the molecular and epigenetic mechanisms that mediate this process in cattle. Compared to other species, cattle have a remarkable ability to accumulate intramuscular fat, partly attributed to the abundance of sources of fatty acids for synthesizing triglycerides. Compared to other adipose depots such as subcutaneous fat, intramuscular fat develops later and grows more slowly. The commitment and differentiation of adipose precursor cells into adipocytes as well as the maturation of adipocytes are crucial steps in intramuscular fat development and growth in cattle. Each of these steps is controlled by various factors, underscoring the complexity of the regulatory network governing adipogenesis in the skeletal muscle. These factors include genetics, epigenetics, nutrition (including maternal nutrition), rumen microbiome, vitamins, hormones, weaning age, slaughter age, slaughter weight, and stress. Many of these factors seem to affect intramuscular fat deposition through the transcriptional or epigenetic regulation of genes directly involved in the development and growth of intramuscular fat. A better understanding of the molecular and cellular mechanisms by which intramuscular fat develops and grows in cattle will help us develop more effective strategies to optimize intramuscular fat deposition in cattle, thereby maximizing the quality and value of beef meat.

Activating Immune Recognition in Pancreatic Ductal Adenocarcinoma via Autophagy Inhibition, MEK Blockade, and CD40 Agonism.

BACKGROUND AND AIMS: Patients with pancreatic ductal adenocarcinoma (PDA) have not yet benefitted from the revolution in cancer immunotherapy due in large part to a dominantly immunosuppressive tumor microenvironment. MEK inhibition combined with autophagy inhibition leads to transient tumor responses in some patients with PDA. We examined the functional effects of combined MEK and autophagy inhibition on the PDA immune microenvironment and the synergy of combined inhibition of MEK and autophagy with CD40 agonism (aCD40) against PDA using immunocompetent model systems. METHODS: We implanted immunologically "cold" murine PDA cells orthotopically in wide type C57BL/6J mice. We administered combinations of inhibitors of MEK1/2, inhibitors of autophagy, and aCD40 and measured anticancer efficacy and immune sequelae using mass cytometry and multiplexed immunofluorescence imaging analysis to characterize the tumor microenvironment. We also used human and mouse PDA cell lines and human macrophages in vitro to perform functional assays to elucidate the cellular effects induced by the treatments. RESULTS: We find that coinhibition of MEK (using cobimetinib) and autophagy (using mefloquine), but not either treatment alone, activates the STING/type I interferon pathway in tumor cells that in turn activates paracrine tumor associated macrophages toward an immunogenic M1-like phenotype. This switch is further augmented by aCD40. Triple therapy (cobimetinib + mefloquine + aCD40) achieved cytotoxic T-cell activation in an immunologically "cold" mouse PDA model, leading to enhanced antitumor immunity. CONCLUSIONS: MEK and autophagy coinhibition coupled with aCD40 invokes immune repolarization and is an attractive therapeutic approach for PDA immunotherapy development.

Evaluation of phase-adjusted interventions for COVID-19 using an improved SEIR model.

A local COVID-19 outbreak with two community clusters occurred in a large industrial city, Shaoxing, China, in December 2021 after serial interventions were imposed. We aimed to understand the reason by analysing the characteristics of the outbreak and evaluating the effects of phase-adjusted interventions. Publicly available data from 7 December 2021 to 25 January 2022 were collected to analyse the epidemiological characteristics of this outbreak. The incubation period was estimated using Hamiltonian Monte Carlo method. A well-fitted extended susceptible-exposed-infectious-recovered model was used to simulate the impact of different interventions under various combination of scenarios. There were 387 SARS-CoV-2-infected cases identified, and 8.3% of them were initially diagnosed as asymptomatic cases. The estimated incubation period was 5.4 (95% CI 5.2-5.7) days for all patients. Strengthened measures of comprehensive quarantine based on tracing led to less infections and a shorter duration of epidemic. With a same period of incubation, comprehensive quarantine was more effective in containing the transmission than other interventions. Our findings reveal an important role of tracing and comprehensive quarantine in blocking community spread when a cluster occurred. Regions with tense resources can adopt home quarantine as a relatively affordable and low-impact intervention measure compared with centralized quarantine.

Do gut microbiome-targeted therapies improve liver function in cirrhotic patients? A systematic review and meta-analysis.

BACKGROUND AND AIM: Microbiome-targeted therapies (MTTs) are considered as promising interventions for cirrhosis, but the impact of gut microbiome modulation on liver function and disease severity has not been fully assessed. We comprehensively evaluated the efficacy of MTTs in patients with liver cirrhosis. METHODS: Data from randomized controlled trials were collected through MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials, and ClinicalTrial.gov from inception to February 20, 2023. Clinical outcomes were pooled and expressed in terms of risk ratios or mean differences (MD). Additional subgroup and sensitivity analyses were performed to validate the robustness of findings. A trial sequential analysis was applied to calculate the required information size and evaluate the credibility of the meta-analysis results. RESULTS: Twenty-one studies with a total of 1699 cirrhotic patients were included for meta-analysis. MTTs were associated with a significant reduction in aspartate aminotransferase (MD, -3.62; 95% CI, -6.59 to -0.65), the risk of hepatic encephalopathy (risk ratio = 0.56, 95% CI: 0.46 to 0.68), model for end-stage liver disease score (MD, -0.90; 95% CI, -1.17 to -0.11), ammonia (MD, -11.86; 95% CI, -16.39 to -7.33), and endotoxin (MD, -0.14; 95% CI, -0.23 to -0.04). The trial sequential analysis yielded reliable results of these outcomes. No effects were observed on the changes of other hepatic function indicators. CONCLUSION: MTTs appeared to be associated with a slowed deterioration in liver cirrhosis, which could provide reference for clinicians in treatment of cirrhotic patients based on their conditions.

Genome-wide identification of enhancers and transcription factors regulating the myogenic differentiation of bovine satellite cells.

BACKGROUND: Satellite cells are the myogenic precursor cells in adult skeletal muscle. The objective of this study was to identify enhancers and transcription factors that regulate gene expression during the differentiation of bovine satellite cells into myotubes. RESULTS: Chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) was performed to identify genomic regions where lysine 27 of H3 histone is acetylated (H3K27ac), i.e., active enhancers, from bovine satellite cells before and during differentiation into myotubes. A total of 19,027 and 47,669 H3K27ac-marked enhancers were consistently identified from two biological replicates of before- and during-differentiation bovine satellite cells, respectively. Of these enhancers, 5882 were specific to before-differentiation, 35,723 to during-differentiation, and 13,199 common to before- and during-differentiation bovine satellite cells. Whereas most of the before- or during-differentiation-specific H3K27ac-marked enhancers were located distally to the transcription start site, the enhancers common to before- and during-differentiation were located both distally and proximally to the transcription start site. The three sets of H3K27ac-marked enhancers were associated with functionally different genes and enriched with different transcription factor binding sites. Specifically, many of the H3K27ac-marked enhancers specific to during-differentiation bovine satellite cells were associated with genes involved in muscle structure and development, and were enriched with binding sites for the MyoD, AP-1, KLF, TEAD, and MEF2 families of transcription factors. A positive role was validated for Fos and FosB, two AP-1 family transcription factors, in the differentiation of bovine satellite cells into myotubes by siRNA-mediated knockdown. CONCLUSIONS: Tens of thousands of H3K27ac-marked active enhancers have been identified from bovine satellite cells before or during differentiation. These enhancers contain binding sites not only for transcription factors whose role in satellite cell differentiation is well known but also for transcription factors whose role in satellite cell differentiation is unknown. These enhancers and transcription factors are valuable resources for understanding the complex mechanism that mediates gene expression during satellite cell differentiation. Because satellite cell differentiation is a key step in skeletal muscle growth, the enhancers, the transcription factors, and their target genes identified in this study are also valuable resources for identifying and interpreting skeletal muscle trait-associated DNA variants in cattle.

Deoxynivalenol interferes with intestinal motility via injuring the contractility of enteric smooth muscle cells: A novel hazard to the gastrointestinal tract by environmental toxins.

Deoxynivalenol (DON) is a prevalent Fusarium mycotoxin, occurs predominantly in the global environment, especially in cereals, animal feed and food commodities. The widespread contamination causes a serious risk to human and animal health. DON usually impairs weight gain, which is presumably from its capacity to reduce feed intake by interfering with intestinal motility. To clarify the role of smooth muscle cells (SMCs) contractility in intestinal motility and growth inhibition caused by DON, twelve weaned piglets were firstly divided into two groups to feed control or Fusarium mycotoxin-contaminated (MC) diet. Results showed that the final body weight, average daily gain and average daily feed intake were significantly reduced in piglets fed the MC diet. Exposure to the MC diet also significantly decreased the thickness of smooth muscle layer and SMCs contractile markers expression (myosin heavy chain 11, smooth muscle actin gamma 2, transgelin, calponin 1) in jejunum and ileum of piglets. Furthermore, oral DON supplementation (3 mg/kg body weight) to mice in six consecutive days could significantly inhibit the upper intestinal transit, impede normal defecation and downregulate SMCs contractile markers expression in small intestine. Finally, we generated a porcine enteric smooth muscle cell line (PISMC), and found that DON could depress its contractility by decreasing PISMC proliferation, migration and contractile markers expression. In conclusion, these findings in vivo and in vitro suggest that DON, as a common environmental toxin, can not only reduce proliferative and motile phenotype, but also decrease contractile apparatus components (contractile markers expression) in SMCs, which in turn influences SMCs contractility and then interferes with intestinal motility and growth performance.

Roles of the proteasome and inhibitor of DNA binding 1 protein in myoblast differentiation.

This study was conducted to further understand the mechanism that controls myoblast differentiation, a key step in skeletal muscle formation. RNA sequencing of primary bovine myoblasts revealed many genes encoding the ubiquitin-proteasome system were up-regulated during myoblast differentiation. This up-regulation was accompanied by increased proteasomal activity. Treating myoblasts with the proteasome-specific inhibitor lactacystin impeded myoblast differentiation. Adenovirus-mediated overexpression of inhibitor of DNA binding 1 (ID1) protein inhibited myoblast differentiation too. Further experiments were conducted to determine whether the proteasome promotes myoblast differentiation by degrading ID1 protein. Both ID1 protein and mRNA expression decreased during myoblast differentiation. However, treating myoblasts with lactacystin reversed the decrease in ID1 protein but not in ID1 mRNA expression. Surprisingly, this reversal was not observed when myoblasts were also treated with the mRNA translation inhibitor cycloheximide. Direct incubation of ID1 protein with proteasomes from myoblasts did not show differentiation stage-associated degradation of ID1 protein. Furthermore, ubiquitinated ID1 protein was not detected in lactacystin-treated myoblasts. Overall, the results of this study suggest that, during myoblast differentiation, the proteasomal activity is up-regulated to further myoblast differentiation and that the increased proteasomal activity improves myoblast differentiation partly by inhibiting the synthesis, not the degradation, of ID1 protein.-Leng, X., Ji, X., Hou, Y., Settlage, R., Jiang, H. Roles of the proteasome and inhibitor of DNA binding 1 protein in myoblast differentiation.

Dual-Emitting N/S-Doped Carbon Dots-Based Ratiometric Fluorescent and Light Scattering Sensor for High Precision Detection of Fe(III) Ions.

Precise and rapid sensing of Fe(III) under concerned facile conditions is important in environmental monitoring. Herein, a facile and label-free ratiometric sensor is constructed for selective determination of Fe(III) ions by coupling second-order scattering (SOS) and fluorescence. We were synthesized fluorescent N, S-doped carbon dots (N/S-CDs) via facile one-step hydrothermal treatment with an intensive fluorescence and a weak SOS signal and high quantum yield (32%). The fluorescence of N/S-CDs was quenched whereas the intensity of SOS was relatively increased by Fe(III) ions due to aggregation-induced fluorescence quenching or enhancement. Based on this effect, a novel fluorescent ratiometric probe with the combined fluorescence and SOS is proposed for the sensing of Fe(III) ions, and with the detection limit of 83 nM and linear range of 0.1-10 µM and 10-40 µM, respectively.

Loss of TIM50 suppresses proliferation and induces apoptosis in breast cancer.

TIM50 is an essential component of TIM23 complex and involved in protein translocating into the inner mitochondrial membrane. Here, we found that TIM50 was increased in breast cancer cells by SILAC. However, its biological functions and molecular mechanisms in breast cancer are poorly understood. To gain insight into the functions of TIM50 in breast cancer, we constructed two stably transfected cell lines and examined TIM50 expression in tissue samples. Our data showed that TIM50 expression was increased in breast cancer. The stable suppression of TIM50 expression through lentivirus-mediated shRNA was shown to inhibit the abilities of cancer cell proliferation and induce apoptosis. What is more, depletion of TIM50 could decrease mitochondrial membrane potential, which may be associated with cell viability. Taken together, our findings reveal a new role for TIM50 in regulating cell proliferation and apoptosis through decreasing mitochondrial membrane potential in breast cancer cell and suggest that TIM50 might be a potential target for controlling breast cancer progression.

Expression of HIV-encoded microRNA-TAR and its inhibitory effect on viral replication in human primary macrophages.

A number of virus-encoded microRNAs have been shown to play important roles in virus replication and virus-host interactions, although the expression and function of miR-TAR-3p derived from the human immunodeficiency virus type 1 (HIV-1) TAR element remain controversial. In this study, miR-TAR-3p was detected in human peripheral blood monocyte-derived macrophages (MDMs) infected by HIV-1. Overexpression of miR-TAR-3p impaired viral replication, while inhibition of miR-TAR-3p enhanced it. Additionally, miR-TAR-3p repressed viral transcription and replication by targeting the TAR element in the HIV-1 5'-LTR in a sequence-specific manner. These results confirm the presence of miR-TAR-3p in HIV-1-infected MDMs and suggest that its function might be used as a mechanism to modulate HIV-1 replication through the expression of a negative regulatory factor.

Short-chain fatty acids enhance adipocyte differentiation in the stromal vascular fraction of porcine adipose tissue.

BACKGROUND: Short-chain fatty acids (SCFAs), including acetate, propionate, and butyrate, are the main products of microbial fermentation in the gut and might mediate some of the effects of gut microbiota and nutrition on development, metabolism, and pathogenesis of obesity and other diseases. OBJECTIVE: The objective of this study was to determine the effects of SCFAs on adipocyte differentiation and the underlying mechanism. METHODS: The stromal vascular fraction (SVF) of the porcine subcutaneous fat was used as the preadipocyte model. Adipocyte differentiation was assessed by Oil Red O staining and gene expression analysis of adipocyte markers. Chromatin immunoprecipitation was used to assess the histone acetylation amounts at the peroxisome proliferator-activated receptor γ (PPARG) and CCAAT/enhancer binding protein α (CEBPA) promoters. RESULTS: Compared with control, propionate and butyrate enhanced the formation of adipocytes by 10-20% and mRNA expression of adipocyte markers by 20-200% in porcine SVF undergoing adipocyte differentiation. Compared with control, short-term treatment of propionate and butyrate enhanced PPARG and CEBPA mRNA expression in porcine SVF by 50-100%. Neither free fatty acid receptor (FFAR) 2 nor FFAR3 mRNA was detectable in porcine SVF before or during differentiation. Neither a cAMP analogue nor an activator of AMP-activated protein kinase (AMPK) affected propionate- or butyrate-enhanced expression of PPARG or CEBPA mRNA. Trichostatin A, a specific inhibitor of histone deacetylases (HDACs), enhanced the formation of adipocytes in porcine SVF by nearly 100% and the expression of PPARG and CEBPA mRNAs by 150% and 50%, respectively. Butyrate increased whereas propionate had no significant effect on histone H3 acetylation at the CEBPA promoter in porcine SVF. CONCLUSIONS: Propionate and butyrate enhance adipocyte differentiation in porcine SVF. These effects are unlikely mediated through FFAR2, FFAR3, cAMP, or AMPK. The effect of butyrate may be partially mediated by its HDAC inhibitory activity, whereas that of propionate is independent of its HDAC inhibitory activity.

Postnatal Growth and Development of the Rumen: Integrating Physiological and Molecular Insights.

The rumen plays an essential role in the physiology and production of agriculturally important ruminants such as cattle. Functions of the rumen include fermentation, absorption, metabolism, and protection. Cattle are, however, not born with a functional rumen, and the rumen undergoes considerable changes in size, histology, physiology, and transcriptome from birth to adulthood. In this review, we discuss these changes in detail, the factors that affect these changes, and the potential molecular and cellular mechanisms that mediate these changes. The introduction of solid feed to the rumen is essential for rumen growth and functional development in post-weaning calves. Increasing evidence suggests that solid feed stimulates rumen growth and functional development through butyric acid and other volatile fatty acids (VFAs) produced by microbial fermentation of feed in the rumen and that VFAs stimulate rumen growth and functional development through hormones such as insulin and insulin-like growth factor I (IGF-I) or through direct actions on energy production, chromatin modification, and gene expression. Given the role of the rumen in ruminant physiology and performance, it is important to further study the cellular, molecular, genomic, and epigenomic mechanisms that control rumen growth and development in postnatal ruminants. A better understanding of these mechanisms could lead to the development of novel strategies to enhance the growth and development of the rumen and thereby the productivity and health of cattle and other agriculturally important ruminants.

Metagenomic and FT-ICR MS insights into the mechanism for the arsenic biogeochemical cycling in groundwater.

Arsenic (As) is a groundwater contaminant of global concern. The degradation of dissolved organic matter (DOM) can provide a reducing environment for As release. However, the interaction of DOM with local microbial communities and how different sources and types of DOM influence the biotransformation of As in aquifers is uncertain. This study used optical spectroscopy, Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS), metagenomics, and structural equation modeling (SEM) to demonstrate the how the biotransformation of As in aquifers is promoted. The results indicated that the DOM in high-As groundwater is dominated by highly unsaturated low-oxygen(O) compounds that are quite humic and stable. Metagenomics analysis indicated Acinetobacter, Pseudoxanthomonas, and Pseudomonas predominate in high-As environments; these genera all contain As detoxification genes and are members of the same phylum (Proteobacteria). SEM analyses indicated the presence of Proteobacteria is positively related to highly unsaturated low-O compounds in the groundwater and conditions that promote arsenite release. The results illustrate how the biogeochemical transformation of As in groundwater systems is affected by DOM from different sources and with different characteristics.

Exploring the Molecular Mechanisms of Huaier on Modulating Metabolic Reprogramming of Hepatocellular Carcinoma: A Study based on Network Pharmacology, Molecular Docking and Bioinformatics.

BACKGROUND: Huaier (Trametes robiniophila Murr), a traditional Chinese medicine, is widely used in China as a complementary and alternative therapy to treat hepatocellular carcinoma (HCC). Past studies have shown that Huaier can arrest the cell cycle, promote apoptosis and inhibit the proliferation of cancer cells. However, how it regulates the metabolism of HCC is still unclear. OBJECTIVE: This study explores the metabolic-related function of Huaier in treating HCC with an in-silico approach. METHODS: A network pharmacology and bioinformatics-based approach was employed to investigate the molecular pathogenesis of metabolic reprogramming in HCC with Huaier. The compounds of Huaier were obtained from public databases. Oral bioavailability and drug likeness were screened using the TCMSP platform. The differential gene expressions between HCC and non-tumor tissue were calculated and used to find the overlap from the targets of Huaier. The enrichment analysis of the overlapped targets by Metascape helped filter out the metabolism-related targets of Huaier in treating HCC. Protein-protein interaction (PPI) network construction and topological screening revealed the hub nodes. The prognosis and clinical correlation of these targets were validated from the cancer genome atlas (TCGA) database, and the interactions between the hub nodes and active ingredients were validated by molecular docking. RESULTS: The results showed that Peroxyergosterol, Daucosterol, and Kaempferol were the primary active compounds of Huaier involved in the metabolic reprogramming of HCC. The top 6 metabolic targets included AKR1C3, CYP1A1, CYP3A4, CYP1A2, CYP17A1, and HSD11B1. The decreased expression of CYP3A4 and increased expression of AKR1C3 were related to the poor overall survival of HCC patients. The molecular docking validated that Peroxyergosterol and Kaempferol exhibited the potential to modulate CYP3A4 and AKR1C3 from a computational perspective. CONCLUSION: This study provided a workflow for understanding the mechanism of Huaier in regulating the metabolic reprogramming of HCC.

Electroacupuncture Facilitates Vascular Normalization by Inhibiting Glyoxalase1 in Endothelial Cells to Attenuate Glycolysis and Angiogenesis in Triple-Negative Breast Cancer.

Recent therapeutic strategies for the treatment of triple-negative breast cancer (TNBC) have shifted the focus from vascular growth factors to endothelial cell metabolism. This study highlights the underexplored therapeutic potential of peri-tumoral electroacupuncture, a globally accepted non-pharmacological intervention for TNBC, and molecular mechanisms. Our study showed that peri-tumoral electroacupuncture effectively reduced the density of microvasculature and enhanced vascular functionality in 4T1 breast cancer xenografts, with optimal effects on day 3 post-acupuncture. The timely integration of peri-tumoral electroacupuncture amplified the anti-tumor efficacy of paclitaxel. Multi-omics analysis revealed Glyoxalase 1 (Glo1) and the associated methylglyoxal-glycolytic pathway as key mediators of electroacupuncture-induced vascular normalization. Peri-tumoral electroacupuncture notably reduced Glo1 expression in the endothelial cells of 4T1 xenografts. Using an in vivo matrigel plug angiogenesis assay, we demonstrated that either Glo1 knockdown or electroacupuncture inhibited angiogenesis. In contrast, Glo1 overexpression increased blood vessel formation. In vitro pharmacological inhibition and genetic knockdown of Glo1 in human umbilical vein endothelial cells inhibited proliferation and promoted apoptosis via downregulating the methylglyoxal-glycolytic pathway. The study using the Glo1-silenced zebrafish model further supported the role of Glo1 in vascular development. This study underscores the pivotal role of Glo1 in peri-tumoral electroacupuncture, spotlighting a promising avenue for enhancing vascular normalization and improving TNBC treatment outcomes.

Enhanced bovine genome annotation through integration of transcriptomics and epi-transcriptomics datasets facilitates genomic biology.

BACKGROUND: The accurate identification of the functional elements in the bovine genome is a fundamental requirement for high-quality analysis of data informing both genome biology and genomic selection. Functional annotation of the bovine genome was performed to identify a more complete catalog of transcript isoforms across bovine tissues. RESULTS: A total of 160,820 unique transcripts (50% protein coding) representing 34,882 unique genes (60% protein coding) were identified across tissues. Among them, 118,563 transcripts (73% of the total) were structurally validated by independent datasets (PacBio isoform sequencing data, Oxford Nanopore Technologies sequencing data, de novo assembled transcripts from RNA sequencing data) and comparison with Ensembl and NCBI gene sets. In addition, all transcripts were supported by extensive data from different technologies such as whole transcriptome termini site sequencing, RNA Annotation and Mapping of Promoters for the Analysis of Gene Expression, chromatin immunoprecipitation sequencing, and assay for transposase-accessible chromatin using sequencing. A large proportion of identified transcripts (69%) were unannotated, of which 86% were produced by annotated genes and 14% by unannotated genes. A median of two 5' untranslated regions were expressed per gene. Around 50% of protein-coding genes in each tissue were bifunctional and transcribed both coding and noncoding isoforms. Furthermore, we identified 3,744 genes that functioned as noncoding genes in fetal tissues but as protein-coding genes in adult tissues. Our new bovine genome annotation extended more than 11,000 annotated gene borders compared to Ensembl or NCBI annotations. The resulting bovine transcriptome was integrated with publicly available quantitative trait loci data to study tissue-tissue interconnection involved in different traits and construct the first bovine trait similarity network. CONCLUSIONS: These validated results show significant improvement over current bovine genome annotations.

Role of glutamate decarboxylase-like protein 1 (GADL1) in taurine biosynthesis.

This manuscript concerns the tissue-specific transcription of mouse and cattle glutamate decarboxylase-like protein 1 (GADL1) and the biochemical activities of human GADL1 recombinant protein. Bioinformatic analysis suggested that GADL1 appears late in evolution, only being found in reptiles, birds, and mammals. RT-PCR determined that GADL1 mRNA is transcribed at high levels in mouse and cattle skeletal muscles and also in mouse kidneys. Substrate screening determined that GADL1, unlike its name implies, has no detectable GAD activity, but it is able to efficiently catalyze decarboxylation of aspartate, cysteine sulfinic acid, and cysteic acid to ß-alanine, hypotaurine, and taurine, respectively. Western blot analysis verified the presence of GADL1 in mouse muscles, kidneys, C2C12 myoblasts, and C2C12 myotubes. Incubation of the supernatant of fresh muscle or kidney extracts with cysteine sulfinic acid resulted in the detection of hypotaurine or taurine in the reaction mixtures, suggesting the possible involvement of GADL1 in taurine biosynthesis. However, when the tissue samples were incubated with aspartate, no ß-alanine production was observed. We proposed several possibilities that might explain the inactivation of ADC activity of GADL1 in tissue protein extracts. Although ß-alanine-producing activity was not detected in the supernatant of tissue protein extracts, its potential role in ß-alanine synthesis cannot be excluded. There are several inhibitors of the ADC activity of GADL1 identified. The discovery of GADL1 biochemical activities, in conjunction with its expression and activities in muscles and kidneys, provides some tangible insight toward establishing its physiological function(s).

Association of growth hormone deficiency with an increased number of preadipocytes in subcutaneous fat.

The inhibitory effect of growth hormone (GH) on adipose tissue growth is well known, but the underlying mechanism is not fully understood. In this study, we determined the possibility that GH inhibits adipose tissue growth by inhibiting adipogenesis, the process of formation of adipocytes from stem cells, in the lit/lit mice. The lit/lit mice are GH deficient because of a spontaneous mutation to the GH releasing hormone receptor (ghrhr) gene, and they have more subcutaneous fat despite being smaller than the lit/+ mice at the same age. We found that cells of the stromal vascular fraction (SVF) of subcutaneous fat from the lit/lit mice had greater adipogenic potential than those from the lit/+ mice, as evidenced by forming greater numbers of lipid droplets-containing adipocytes and having greater expression of adipocyte marker genes during induced adipocyte differentiation in culture. However, addition of GH to the culture did not reverse the superior adipogenic potential of subcutaneous SVF from the lit/lit mice. Through florescence-activated cell sorting and quantification of mRNAs of preadipocyte markers, including CD34, CD29, Sca-1, CD24, Pref-1, and PPARγ, we found that subcutaneous SVF from the lit/lit mice contained more preadipocytes than that from the lit/+ mice. These results support the notion that GH inhibits adipose tissue growth in mice at least in part by inhibiting adipogenesis. Furthermore, these results suggest that GH inhibits adipogenesis in mice not by inhibiting the terminal differentiation of preadipocytes into adipocytes, rather by inhibiting the formation of preadipocytes from stem cells or the recruitment of stem cells to the fat depot.

Autoantibodies against type I interferons in COVID-19 infection: A systematic review and meta-analysis.

OBJECTIVES: In this study, we aimed to study the rate of autoantibodies against type I interferons (IFNs) in patients with COVID-19 and analyze its dependence on severity of infection and some other variables. METHODS: A systemic review with the search terms: "COVID-19" or "SARS-CoV-2" and "autoantibodies" or "autoantibody" and "IFN" or "interferon" for the period 20 December 2019 to 15 August 2022 was carried out using PubMed, Embase, Cochrane, and Web of Science. R 4.2.1 software was used for meta-analysis of the published results. Pooled risk ratios and 95% confidence intervals (CIs) were calculated. RESULTS: We identified eight studies involving 7729 patients, of whom 5097 (66%) had severe COVID-19 and 2632 (34%) had mild or moderate symptoms. The positive rate of anti-type-I-IFN-autoantibodies in the total dataset was 5% (95% CI, 3-8%), but reached 10% (95% CI, 7-14%) in those with severe infection. The most common subtypes were anti-IFN-α (89%) and anti-IFN-ω (77%). The overall prevalence in male patients was 5% (95% CI, 4-6%), and in female patients 2% (95% CI, 1-3%). CONCLUSION: Severe COVID-19 is associated with high rates of autoantibodies against type-I-IFN and more so in male than female patients.