Epigenetic regulation in insects may have effects on diverse biological processes. Here we survey the methylome of a model insect, the silkworm Bombyx mori, at single-base resolution using Illumina high-throughput bisulfite sequencing (MethylC-Seq). We conservatively estimate that 0.11% of genomic cytosines are methylcytosines, all of which probably occur in CG dinucleotides. CG methylation is substantially enriched in gene bodies and is positively correlated with gene expression levels, suggesting it has a positive role in gene transcription. We find that transposable elements, promoters and ribosomal DNAs are hypomethylated, but in contrast, genomic loci matching small RNAs in gene bodies are densely methylated. This work contributes to our understanding of epigenetics in insects, and in contrast to previous studies of the highly methylated genomes of Arabidopsis and human, demonstrates a strategy for sequencing the epigenomes of organisms such as insects that have low levels of methylation.

Analysis across the genome of patterns of DNA methylation reveals a rich landscape of allele-specific epigenetic modification and consequent effects on allele-specific gene expression.

BackgroundDNA methylation plays important biological roles in plants and animals. To examine the rice genomic methylation landscape and assess its functional significance, we generated single-base resolution DNA methylome maps for Asian cultivated rice Oryza sativa ssp. japonica, indica and their wild relatives, Oryza rufipogon and Oryza nivara.ResultsThe overall methylation level of rice genomes is four times higher than that of Arabidopsis. Consistent with the results reported for Arabidopsis, methylation in promoters represses gene expression while gene-body methylation generally appears to be positively associated with gene expression. Interestingly, we discovered that methylation in gene transcriptional termination regions (TTRs) can significantly repress gene expression, and the effect is even stronger than that of promoter methylation. Through integrated analysis of genomic, DNA methylomic and transcriptomic differences between cultivated and wild rice, we found that primary DNA sequence divergence is the major determinant of methylational differences at the whole genome level, but DNA methylational difference alone can only account for limited gene expression variation between the cultivated and wild rice. Furthermore, we identified a number of genes with significant difference in methylation level between the wild and cultivated rice.ConclusionsThe single-base resolution methylomes of rice obtained in this study have not only broadened our understanding of the mechanism and function of DNA methylation in plant genomes, but also provided valuable data for future studies of rice epigenetics and the epigenetic differentiation between wild and cultivated rice.

The landscape of genetic alterations in lung adenocarcinoma derived from Asian patients is largely uncharacterized. Here we present an integrated genomic and transcriptomic analysis of 335 primary lung adenocarcinomas and 35 corresponding lymph node metastases from Chinese patients. Altogether 13 significantly mutated genes are identified, including the most commonly mutated gene TP53 and novel mutation targets such as RHPN2, GLI3 and MRC2. TP53 mutations are furthermore significantly enriched in tumours from patients harbouring metastases. Genes regulating cytoskeleton remodelling processes are also frequently altered, especially in metastatic samples, of which the high expression level of IQGAP3 is identified as a marker for poor prognosis. Our study represents the first large-scale sequencing effort on lung adenocarcinoma in Asian patients and provides a comprehensive mutational landscape for both primary and metastatic tumours. This may thus form a basis for personalized medical care and shed light on the molecular pathogenesis of metastatic lung adenocarcinoma.

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People of East Asian ethnicity have a different prevalence of and show unique clinical characteristics and tumor histology of oncogenic mutations. However, only limited studies have explored the landscape of genomic alterations in lung adenocarcinoma derived from Asian patients thus far. In this single‐center study, with an aim to elucidate the mutational profile of lung cancer in people of Chinese ethnicity and to use the obtained information to guide decision‐making for treatment, we employed a well‐validated assay to perform comprehensive genomic characterization of tumor specimens from 306 Chinese lung cancer patients. A total of 845 individual genomic alterations were found in 145 tumor‐related genes with a median of 2.8 alterations (range: 1–18) per sample. The most frequently mutated genes were EGFR (46.7%), TP53 (21.2%), ALK (12.1%; 8.8% of mutation and 3.3% of rearrangement) and KRAS (10.1%). Upon comparison with the Cancer Genome Atlas dataset, we found that EGFR was mutated at a much higher frequency in our cohort than in Caucasians, whereas KRAS was only found in 10.1% of our Chinese patients. Clinically relevant genomic alterations were identified in 185 (60.5%) patients, including 50% in adenocarcinoma patients and 14% in squamous cell carcinoma patients. Our findings suggest that the Asian ethnicity is significantly different from the Caucasian ethnicity with regard to the presence of somatic driver mutations. Furthermore, we showed that the use of a comprehensive genotyping approach could help identify actionable genomic alterations that have potential impact on therapeutic decisions.

Objective: To evaluate the diagnostic performance of donor-derived plasma cell-free DNA (cfDNA) in discriminating antibody-mediated rejection (ABMR) or de novo donorspecific antibodies (DSA) without histological lesions in kidney allograft recipients. Methods:In this prospective single center observational study, we enrolled kidney allograft recipients between November, 2016 and September, 2017 at the First Affiliated Hospital of Sun Yat-sen University. Kidney allograft recipients with ABMR, de novo DSA but no histological lesions or negative DSA, and stable renal function were included. The plasma cfDNA fraction was measured using a targeted, single nucleotide polymorphism (SNP)-based assay. Pathological diagnosis was made according to the 2015 Banff Kidney Rejection Classification. The area under the ROC curve (AUC-ROC) was determined using the bootstrapping method to estimate median and 95% confidence interval (95% CI). The sensitivity, specificity and Youden index, positive predictive value (PPV), and negative predictive value (NPV) were calculated for specific cfDNA fractions.Results: Totally 37 consecutive patients received kidney allografts, including 18 recipients in the ABMR group and 19 recipients in the stable allograft group (7 DSApositive and 12 DSA-negative). All patients in the ABMR group were DSA positive and 7 patients in the stable group were DSA positive but had no pathologically proven ABMR. The median donor-derived plasma cfDNA fraction was 2.4% (Q1 1.52% -Q3 3.70%) in the ABMR group, and was significantly higher than that of the stable group (0.65%, Q1 Frontiers in Immunology | www.frontiersin.org February 2020 | Volume 11 | Article 342 Zhang et al.Donor-Derived cfDNA in Kidney ABMR 0.57% -Q3 0.97%; P < 0.001), but comparable with that of the DSA-positive patients in the stable allograft group (P = 0.074). The AUC-ROC of cfDNA was 0.90 (95% CI, 0.79-0.98). When a cfDNA threshold of 1% was chosen, it had a sensitivity of 88.9% and a specificity of 73.7%. The PPV was 76.2% and the NPV was 87.5%.Conclusion: Donor-derived plasma cfDNA fraction increased in kidney allograft recipients with ABMR. Detection of donor-derived plasma cfDNA fraction may contribute to the discrimination between ABMR and stable renal allograft function and may aid early recognition of earlier stage antibody-mediated injury.

Molecular profiling of lung cancer has become essential for prediction of an individual’s response to targeted therapies. Next-generation sequencing (NGS) is a promising technique for routine diagnostics, but has not been sufficiently evaluated in terms of feasibility, reliability, cost and capacity with routine diagnostic formalin-fixed, paraffin-embedded (FFPE) materials. Here, we report the validation and application of a test based on Ion Proton technology for the rapid characterisation of single nucleotide variations (SNVs), short insertions and deletions (InDels), copy number variations (CNVs), and gene rearrangements in 145 genes with FFPE clinical specimens. The validation study, using 61 previously profiled clinical tumour samples, showed a concordance rate of 100% between results obtained by NGS and conventional test platforms. Analysis of tumour cell lines indicated reliable mutation detection in samples with 5% tumour content. Furthermore, application of the panel to 58 clinical cases, identified at least one actionable mutation in 43 cases, 1.4 times the number of actionable alterations detected by current diagnostic tests. We demonstrated that targeted NGS is a cost-effective and rapid platform to detect multiple mutations simultaneously in various genes with high reproducibility and sensitivity.