Somatic embryogenesis is an important tissue culture technique that sometimes leads to phenotypic variation via genetic and/or epigenetic changes. To understand the genomic and epigenomic impacts of somatic embryogenesis, we characterized soybean (Glycine max) epigenomes sampled from embryos at 10 different stages ranging from 6 weeks to 13 years of continuous culture. We identified genome-wide increases in DNA methylation from cultured samples, especially at CHH sites. The hypermethylation almost exclusively occurred in regions previously possessing non-CG methylation and was accompanied by increases in the expression of genes encoding the RNA-directed DNA methylation (RdDM) machinery. The epigenomic changes were similar between somatic and zygotic embryogenesis. Following the initial global wave of hypermethylation, rare decay events of maintenance methylation were observed, and the extent of the decay increased with time in culture. These losses in DNA methylation were accompanied by downregulation of genes encoding the RdDM machinery and transcriptome reprogramming reminiscent of transcriptomes during late-stage seed development. These results reveal a process for reinforcing already silenced regions to maintain genome integrity during somatic embryogenesis over the short term, which eventually decays at certain loci over longer time scales.

To understand the variation in genomic patterning of DNA methylation we compared methylomes of 34 diverse angiosperm species. By analyzing whole-genome bisulfite sequencing data in a phylogenetic context it becomes clear that there is extensive variation throughout angiosperms in gene body DNA methylation, euchromatic silencing of transposons and repeats, as well as silencing of heterochromatic transposons. The Brassicaceae have reduced CHG methylation levels and also reduced or loss of CG gene body methylation. The Poaceae are characterized by a lack or reduction of heterochromatic CHH methylation and enrichment of CHH methylation in genic regions. Reduced CHH methylation levels are found in clonally propagated species, suggesting that these methods of propagation may alter the epigenomic landscape over time. These results show that DNA methylation patterns are broadly a reflection of the evolutionary and life histories of plant species.

Preclinical studies have shown that interleukin (IL)-1β blockade can modulate the tumor microenvironment (TME) to activate antitumor immunity and, in combination with immune checkpoint inhibitors (ICIs), prevent cancer growth. Our study investigates if immune biomarkers in the TME affect outcomes in patients with non–small cell lung cancer (NSCLC) treated with the IL-1β inhibitor canakinumab plus an ICI-based therapy and describes canakinumab effects on the TMEs in these patients. Exploratory analyses were conducted in two prospective trials evaluating canakinumab combined with pembrolizumab-based regimens in patients with NSCLC: CANOPY-1 (first-line setting) and CANOPY-N (neoadjuvant setting). Immunohistochemistry (IHC) and transcriptomic analyses were performed on baseline tumor samples from CANOPY-1, and IHC and multiplex immunofluorescence analyses were performed on baseline and posttreatment tumor samples from CANOPY-N. Associations with clinical outcomes were evaluated. In CANOPY-1, in patients with low levels of T-cell infiltration in the tumor, the addition of canakinumab to a pembrolizumab-based regimen was associated with progression-free and overall survival improvements. Low levels of T-cell infiltration were associated with an immunosuppressive gene expression phenotype, supporting the role of low T-cell infiltration as a surrogate of an overall immunosuppressive TME. In CANOPY-N, a reduction in immunosuppressive cells in the TME was observed following canakinumab and pembrolizumab treatment. Our exploratory biomarker analyses from the CANOPY-1 and CANOPY-N trials suggest that IL-1β blockade may shift the TME toward an immune-activated status and that patients with immunosuppressive TME features could benefit from the addition of canakinumab to an ICI-based treatment. Significance: Patients with NSCLC with immunosuppressive tumor features and low T-cell infiltration derive less benefit from ICI-based treatment. Biomarker analyses presented here suggest that these patients may benefit from the addition of anti–IL-1β therapy to their treatment.

Background: The evolution of gene body methylation (gbM), its origins and its functional consequences are poorly understood. By pairing the largest collection of transcriptomes (>1000) and methylomes (77) across Viridiplantae we provide novel insights into the evolution of gbM and its relationship to CHROMOMETHYLASE (CMT) proteins. Results: CMTs are evolutionary conserved DNA methyltransferases in Viridiplantae. Duplication events gave rise to what are now referred to as CMT1, 2 and 3. Independent losses of CMT1, 2 and 3 in eudicots, CMT2 and ZMET in monocots and monocots/commelinids, variation in copy number and non-neutral evolution suggests overlapping or fluid functional evolution of this gene family. DNA methylation within genes is widespread and is found in all major taxonomic groups of Viridiplantae investigated. Genes enriched with methylated CGs (mCG) were also identified in species sister to angiosperms. The proportion of genes and DNA methylation patterns associated with gbM are restricted to angiosperms with a functional CMT3 or ortholog. However, mCG-enriched genes in the gymnosperm Pinus taeda shared some similarities with gbM genes in Amborella trichopoda . Additionally, gymnosperms and ferns share a CMT homolog closely related to CMT2 and 3. Hence, the dependency of gbM on a CMT most likely extends to all angiosperms and possibly gymnosperms and ferns. Conclusions: The resulting gene family phylogeny of CMT transcripts from the most diverse sampling of plants to date redefines our understanding of CMT evolution and its evolutionary consequences on DNA methylation. Future, functional tests of homologous and paralogous CMTs will uncover novel roles and consequences to the epigenome.

Immune checkpoint inhibitors (ICI) have demonstrated clinical efficacy in non–small cell lung cancer (NSCLC), and extensive research has been conducted to explore biomarkers predictive of ICI response. However, the impact of ICI on the tumor and tumor microenvironment at primary and acquired resistance states is understudied due to the difficulty of collecting tissue biopsies at disease progression. In this study, we leveraged clinical and real-world data to study ICI resistance. Data used in this work consist of treatment outcome information and tissue RNA sequencing data from advanced-stage NSCLC cohorts from three sources: the Tempus real-world evidence database; CANOPY-1 (NCT03631199), a phase III clinical trial in first-line NSCLC; and Stand Up To Cancer (SU2C) publication. Our results indicate higher IFNγ and T-cell exhaustion in patients’ tumors at acquired resistance and low levels of B-cell and dendritic cell expression at primary resistance. The lower B-cell and dendritic cell levels may be primarily driven by prior treatment with a platinum-based chemotherapy regimen. Baseline transcriptomics data additionally suggest that innate immune cells may play an antitumor role in PD-L1<1% patients, whereas IFNγ and T-cell inflammation are more predictive of ICI treatment outcomes in PD-L1≥1% patients. Our study suggests a clear divergence of the tumor microenvironment in patients with primary versus acquired resistance and a potential role of myeloid cells in the PD-L1<1% population. These findings shed light on potential next-generation therapies to overcome ICI resistance. Significance: ICI benefits patients with NSCLC, but resistance remains common. Our research highlights differences in tumor environments between primary and acquired resistance after ICI treatment, emphasizing distinct post-therapy approaches. Findings also suggest myeloid cells as key players in PD-L1–negative cases, guiding future treatment strategies to overcome resistance and improve outcomes.