Genomic and oncogenic preference of HBV integration in hepatocellular carcinoma

Hepatitis B virus (HBV) can integrate into the human genome, contributing to genomic instability and hepatocarcinogenesis. Here by conducting high-throughput viral integration detection and RNA sequencing, we identify 4,225 HBV integration events in tumour and adjacent non-tumour samples from 426 patients with HCC. We show that HBV is prone to integrate into rare fragile sites and functional genomic regions including CpG islands. We observe a distinct pattern in the preferential sites of HBV integration between tumour and non-tumour tissues. HBV insertional sites are significantly enriched in the proximity of telomeres in tumours. Recurrent HBV target genes are identified with few that overlap. The overall HBV integration frequency is much higher in tumour genomes of males than in females, with a significant enrichment of integration into chromosome 17. Furthermore, a cirrhosis-dependent HBV integration pattern is observed, affecting distinct targeted genes. Our data suggest that HBV integration has a high potential to drive oncogenic transformation

Restraint stress promotes nonalcoholic steatohepatitis by regulating the farnesoid X receptor/NLRP3 signaling pathway

Psychological stress promotes nonalcoholic steatohepatitis (NASH) development. However, the pathogenesis of psychological stress-induced NASH remains unclear. This study aims to explore the underlying mechanism of restraint stress-induced NASH, which mimics psychological stress, and to discover potential NASH candidates. Methionine choline deficient diet- and high fat diet-induced hepatosteatotic mice are subjected to restraint stress to induce NASH. The mice are administrated with Xiaoyaosan granules, NOD-like receptor family pyrin domain containing 3 (NLRP3) inhibitors, farnesoid X receptor (FXR) agonists, or macrophage scavengers. Pathological changes and NLRP3 signaling in the liver are determined. These results demonstrate that restraint stress promotes hepatic inflammation and fibrosis in hepatosteatotic mice. Restraint stress increases the expressions of NLRP3, Caspase-1, Gasdermin D, interleukin-1β, cholesterol 7α-hydroxylase, and sterol 12α-hydroxylase and decreases the expression of FXR in NASH mice. Xiaoyaosan granules reverse hepatic inflammation and fibrosis and target FXR and NLRP3 signals. In addition, inhibition of NLRP3 reduces the NLRP3 inflammasome and liver damage in mice with restraint stress-induced NASH. Elimination of macrophages and activation of FXR also attenuate inflammation and fibrosis by inhibiting NLRP3 signaling. However, NLRP3 inhibitors or macrophage scavengers fail to affect the expression of FXR. In conclusion, restraint stress promotes NASH-related inflammation and fibrosis by regulating the FXR/NLRP3 signaling pathway. Xiaoyaosan granules, NLRP3 inhibitors, FXR agonists, and macrophage scavengers are potential candidates for the treatment of psychological stress-related NASH

The Reduced Plastid-Encoded Polymerase-Dependent Plastid Gene Expression Leads to the Delayed Greening of the <i>Arabidopsis fln2</i> Mutant

<div><p>In <i>Arabidopsis</i> leaf coloration mutants, the delayed greening phenomenon is common. Nonetheless, the mechanism remains largely elusive. Here, a delayed greening mutant <i>fln2–4</i> of FLN2 (Fructokinase-Like Protein2) was studied. FLN2 is one component of Transcriptionally Active Chromosome (TAC) complex which is thought to contain the complete plastid-encoded polymerase (PEP). <i>fln2–4</i> displayed albino phenotype on medium without sucrose. The PEP-dependent plastid gene expression and chloroplast development were inhibited in <i>fln2–4</i>. Besides interacting with thioredoxin z (TRX z), we identified that FLN2 interacted with another two members of TAC complex in yeast including its homologous protein FLN1 (Fructokinase-Like Protein1) and pTAC5. This indicates that FLN2 functions in regulation of PEP activity associated with these TAC components. <i>fln2–4</i> exhibited delayed greening on sucrose-containing medium. Comparison of the PEP-dependent gene expression among two complete albino mutants (<i>trx z</i> and <i>ptac14</i>), two yellow mutants (<i>ecb2–2</i> and <i>ys1</i>) and the <i>fln2–4</i> showed that <i>fln2–4</i> remains partial PEP activity. FLN2 and FLN1 are the target proteins of TRX z involved in affecting the PEP activity. Together with the data that FLN1 could interact with itself in yeast, FLN1 may form a homodimer to replace FLN1–FLN2 as the TRX z target in redox pathway for maintaining partial PEP activity in <i>fln2–4</i>. We proposed the partial PEP activity in the <i>fln2</i> mutant allowed plastids to develop into fully functional chloroplasts when exogenous sucrose was supplied, and finally the mutants exhibited green phenotype.</p></div

Changes in the transcript levels of PEP-dependent genes during the greening process of <i>fln2–4</i> mutant.

<p>The expression levels of <i>rbcL</i>, <i>psbA</i> and <i>psbB</i> genes in 7-day-old and 14-day-old <i>fln2–4</i> mutants were determined by Northern blot as compared with WT, respectively. The experimental WT and <i>fln2–4</i> seedlings were grown on sucrose-containing MS medium.</p

Chlorophyll accumulation in the WT and <i>fln2–4</i> mutants.

*<p>Averages ± standard deviations of chlorophyll (Chl) concentrations for 3 independent measurements. FW: fresh weight.</p

Comparison of PEP-dependent plastid gene expression between <i>fln2–4</i> and four leaf coloration mutants.

<p>(A) The phenotypes of WT and four leaf coloration mutants grown with or without sucrose. (B) qRT-PCR analysis the transcript levels of four PEP-dependent plastid genes in <i>fln2–4</i> seedlings and other four leaf coloration mutants grown on MS medium without sucrose for 7 days. These PEP-dependent genes refer to <i>psbA</i>, <i>psbB</i>, <i>psaB</i>, <i>petD</i>. Expression levels are presented as the percentage relative to WT. Data are means ± SD (n = 3). (C) The accumulation of <i>psbA</i> and <i>psbB</i> transcripts detected by Northern blot.</p

Expression analysis of the plastid encoded genes in <i>fln2–4</i> seedlings.

<p>Northern blot was performed to detect the plastid gene transcript levels in the 7-day-old <i>fln2–4</i> seedlings and WT grown on MS medium without sucrose. Three classes of genes were examined, <i>psbA, psbB</i>, and <i>rbcL</i> were selected as PEP-dependent genes, <i>clpP</i> and <i>rrn16</i> were selected as PEP- and NEP-dependent genes, <i>accD</i> and <i>rpoA</i> were selected as NEP-dependent genes.</p

Continuous exposure to isoprenaline reduced myotube size by delaying myoblast differentiation and fusion through the NFAT-MEF2C signaling pathway

Abstract We aimed to explore whether superfluous sympathetic activity affects myoblast differentiation, fusion, and myofiber types using a continuous single-dose isoprenaline exposure model in vitro and to further confirm the role of distinct NFATs in ISO-mediated effects. Compared with delivery of single and interval single, continuous single-dose ISO most obviously diminished myotube size while postponing myoblast differentiation/fusion in a time- and dose-dependent pattern, accompanied by an apparent decrease in nuclear NFATc1/c2 levels and a slight increase in nuclear NFATc3/c4 levels. Overexpression of NFATc1 or NFATc2, particularly NFATc1, markedly abolished the inhibitory effects of ISO on myoblast differentiation/fusion, myotube size and Myh7 expression, which was attributed to a remarkable increase in the nuclear NFATc1/c2 levels and a reduction in the nuclear NFATc4 levels and the associated increase in the numbers of MyoG and MEF2C positive nuclei within more than 3 nuclei myotubes, especially in MEF2C. Moreover, knockdown of NFATc3 by shRNA did not alter the inhibitory effect of ISO on myoblast differentiation/fusion or myotube size but partially recovered the expression of Myh7, which was related to the slightly increased nuclear levels of NFATc1/c2, MyoG and MEF2C. Knockdown of NFATc4 by shRNA prominently increased the number of MyHC +, MyoG or MEF2C + myoblast cells with 1 ~ 2 nuclei, causing fewer numbers and smaller myotube sizes. However, NFATc4 knockdown further deteriorated the effects of ISO on myoblast fusion and myotube size, with more than 5 nuclei and Myh1/2/4 expression, which was associated with a decrease in nuclear NFATc2/c3 levels. Therefore, ISO inhibited myoblast differentiation/fusion and myotube size through the NFAT-MyoG-MEF2C signaling pathway