Plant leaves, harvesting light energy and fixing CO 2 , are a major source of foods on the earth. Leaves undergo developmental and physiological shifts during their lifespan, ending with senescence and death. We characterized the key regulatory features of the leaf transcriptome during aging by analyzing total-and small-RNA transcriptomes throughout the lifespan of Arabidopsis (Arabidopsis thaliana) leaves at multidimensions, including age, RNA-type, and organelle. Intriguingly, senescing leaves showed more coordinated temporal changes in transcriptomes than growing leaves, with sophisticated regulatory networks comprising transcription factors and diverse small regulatory RNAs. The chloroplast transcriptome, but not the mitochondrial transcriptome, showed major changes during leaf aging, with a strongly shared expression pattern of nuclear transcripts encoding chloroplast-targeted proteins. Thus, unlike animal aging, leaf senescence proceeds with tight temporal and distinct interorganellar coordination of various transcriptomes that would be critical for the highly regulated degeneration and nutrient recycling contributing to plant fitness and productivity.Most organisms undergo age-dependent developmental changes during their lifespans. The timely decision of developmental changes during the lifespan is a critical evolutionary characteristic that maximizes fitness in a given ecological setting (Leopold, 1961;Fenner, 1998;Samach and Coupland, 2000). Plants use unique developmental strategies throughout their lifespans as opposed to animals. In plants, most organs are formed postnatally from sets of stem cells in the seed. In addition, plants are sessile and cope with encountering environments physiologically, rather than behaviorally. Thus, they have developed highly plastic and interactive developmental programs to incorporate environmental changes into their developmental decisions (Pigliucci, 1998;Sultan, 2000).The leaf is an organ that characterizes the fundamental aspects of plants. Leaves harvest light energy, fix CO 2 to produce carbohydrates, and, as primary producers in our ecosystem, serve as a major food source on the earth. Leaves undergo a series of developmental and physiological shifts during their lifespans. A leaf is initially formed as a leaf primordium derived from the stem cells at the shoot apical meristem and develops into a photosynthetic organ through biogenesis processes involving cell division, differentiation, and expansion (Tsukaya, 2013). In the later stages of their lifespans, leaves undergo organ-level senescence and eventually death. Organlevel senescence in plants involves postmitotic senescence and is a term used similarly as "aging" in animals. During the senescence stage, leaf cells undergo dramatic shifts in physiology from biogenesis to the sequential 1 This research was supported by the Institute for Basic Science (IBS-R013-D1 and IBS-R013-G1), the DGIST R&D Program (2014010043, 2015010004, 2015010011, 20150100012, and 15-01-HRLA-01), Basic Science Research Program (2010-0...

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Supplementary data are available at Bioinformatics online.

Recently, MET exon 14 deletion (METex14del) has been postulated to be one potential mechanism for MET protein overexpression. We screened for the presence of METex14del transcript by multiplexed fusion transcript analysis using nCounter assay followed by confirmation with quantitative reverse transcription PCR with correlation to MET protein expression by immunohistochemistry (IHC) and MET amplification by fluorescence in situ hybridization (FISH). We extracted RNAs from 230 patients enrolled onto the prospective molecular profiling clinical trial (NEXT-1) (NCT02141152) between November 2013 and August 2014. Thirteen METex14del cases were identified including 3 gastric cancer, 4 colon cancer, 5 non-small cell lung cancer, and one adenocarcinoma of unknown primary. Of these 13 METex14del cases, 11 were MET IHC 3+ and 2 were 2+. Only one out of the 13 METex14del cases was MET amplified (MET/CEP ratio > 2.0). Growths of two (gastric, colon) METex14del+ patient tumor derived cell lines were profoundly inhibited by both MET tyrosine kinase inhibitors and a monoclonal antibody targeting MET. In conclusion, METex14del is a unique molecular aberration present in gastrointestinal (GI) malignancies corresponding with overexpression of MET protein but rarely with MET amplification. Substantial growth inhibition of METex14del+ patient tumor derived cell lines by several MET targeting drugs strongly suggests METex14del is a potential actionable driver mutation in GI malignancies.

BackgroundIn this study, we established patient-derived tumor cell (PDC) models using tissues collected from patients with metastatic cancer and assessed whether these models could be used as a tool for genome-based cancer treatment.MethodsPDCs were isolated and cultured from malignant effusions including ascites and pleural fluid. Pathological examination, immunohistochemical analysis, and genomic profiling were performed to compare the histological and genomic features of primary tumors, PDCs. An exploratory gene expression profiling assay was performed to further characterize PDCs.ResultsFrom January 2012 to May 2013, 176 samples from patients with metastatic cancer were collected. PDC models were successfully established in 130 (73.6%) samples. The median time from specimen collection to passage 1 (P1) was 3 weeks (range, 0.5–4 weeks), while that from P1 to P2 was 2.5 weeks (range, 0.5–5 weeks). Sixteen paired samples of genomic alterations were highly concordant between each primary tumor and progeny PDCs, with an average variant allele frequency (VAF) correlation of 0.878. We compared genomic profiles of the primary tumor (P0), P1 cells, P2 cells, and patient-derived xenografts (PDXs) derived from P2 cells and found that three samples (P0, P1, and P2 cells) were highly correlated (0.99–1.00). Moreover, PDXs showed more than 100 variants, with correlations of only 0.6–0.8 for the other samples. Drug responses of PDCs were reflective of the clinical response to targeted agents in selected patient PDC lines.Conclusion(s)Our results provided evidence that our PDC model was a promising model for preclinical experiments and closely resembled the patient tumor genome and clinical response.

Single-cell transcriptomic profiles analysis has proposed new insights for understanding the behavior of human gastric cancer (GC). GC offers a unique model of intratumoral heterogeneity. However, the specific classes of cells involved in carcinogenetic passage, and the tumor microenvironment of stromal cells was poorly understood. We characterized the heterogeneous cell population of precancerous lesions and gastric cancer at the single-cell resolution by RNA sequencing. We identified 10 gastric cell subtypes and showed the intestinal and diffuse-type cancer were characterized by different cell population. We found that the intestinal and diffuse-type cancer cells have the differential metaplastic cell lineages: intestinal-type cancer cells differentiated along the intestinal metaplasia lineage while diffuse-type cancer cells resemble de novo pathway. We observed an enriched CCND1 mutation in premalignant disease state and discovered cancer-associated fibroblast cells harboring pro-stemness properties. In particular, tumor cells could be categorized into previously proposed molecular subtypes and harbored specific subtype of malignant cell with high expression level of epithelial-myofibroblast transition which was correlated with poor clinical prognosis. In addition to intratumoral heterogeneity, the analysis revealed different cellular lineages were responsible for potential carcinogenetic pathways. Single-cell transcriptomes analysis of gastric pre-cancerous lesions and cancer may provide insights for understanding GC cell behavior, suggesting potential targets for the diagnosis and treatment of GC.

Tumor infiltrating lymphocytes (TIL) in Epstein-Barr virus (EBV)-associated/microsatellite-unstable (MSI) gastric carcinomas (GC) constitute immune-active principal cellular components of tumor microenvironment and contribute to better prognosis. With the remarkable success of cancer immunotherapies, there is an urgent need for a comprehensive understanding of tumor-immune interactions in patients with GC in the context of host immune response. To identify GC subtype-specific immune response gene set, we tested differentially expressed genes for MSI and EBV+ GC subtypes in randomly selected test set (n = 278) in merged ACRG-SMC microarray and TCGA RNA sequencing data set. We identified Host ImmunE Response index (HIERÏ) consisting of 29 immune genes classifying GC patients into robust 3 groups with prognostic significance. Immune-high cluster 1 was enriched with PD-L1/EBV+/MSI/TIL with the best clinical outcome while immune-low cluster 3 displayed worst outcome and exemplified with PD-L1/EBV-/MSS. The results were validated in the same cohort (n = 279) and independent cohort (n = 181) with RNA from formalin-fixed paraffin-embedded (FFPE) tissue. Unexpectedly, nearly half of GC in cluster 1 were EBV-/MSS and 10% of cluster 3 GC were EBV+/MSI GC patients, suggesting that in addition to EBV+/MSI GC subtypes, EBV-/MSS subtype also constitutes almost half of high immune cluster and would be a good candidate for immune checkpoint inhibitor therapy. In contrary, almost 10% of EBV+/MSI GC patients may not respond to immune checkpoint inhibitor therapy. Thus, our HIERÏ gene signature demonstrates the potential to subclassify tumor immunity levels, predict prognosis and help immunotherapeutic decisions.

Deamination of nucleotides causes C:G>T:A changes in formalin-fixed, paraffin-embedded (FFPE) tissue samples and produces false positives during next-generation sequencing (NGS). Uracil DNA glycosylase (UDG) helps eliminate this issue, but the effect of UDG in different tissue preparation conditions has not been rigorously studied. To investigate whether UDG can reduce false-positive single-nucleotide variant (SNV) calls, we used tumor and normal tissues from gastric adenocarcinoma patients prepared using different fixation times and pH conditions. FFPE tumor blocks >10 years were also evaluated for the comparison. We performed semiconductor-based NGS to evaluate nucleotide changes and used UDG to test deamination-related effects. Sequencing quality parameters mildly worsened with prolonged fixation time, acidic pH, and delayed fixation. SNV calls and C:G>T:A changes increased after >48 hours of fixation. In both recently prepared and old FFPE tissue blocks, UDG treatment reduced deamination-induced nucleotide changes. In the recently prepared samples, both high-quality SNVs and mean target coverage were remarkably increased on treatment with UDG. However, the quality of NGS results from old-age samples varied irrespective of UDG treatment. In conclusion, based on our findings, we believe that when performing NGS on recently embedded blocks, it is important to consider that certain poorly fixed samples may be at the risk of being deaminated, which can be corrected with UDG treatment.