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The endosymbiosis between cnidarians and photosynthetic dinoflagellates of the Symbiodiniaceae family forms the foundation of coral reef ecosystems. Prolonged environmental shifts can disrupt the cnidarian–Symbiodiniaceae partnership, triggering dysbiosis and coral bleaching and ultimately resulting in coral starvation, mortality, and the collapse of reef ecosystems. Despite its significance, critical gaps remain in our understanding of the cellular mechanisms governing symbiosis and dysbiosis. Innate immune genes and pathways are highly conserved across the Metazoa, including in cnidarians. Among these is NADPH oxidase (NOX), a key enzyme responsible for generating reactive oxygen species (ROS), primarily for microbial degradation within phagolysosomes. In this study, we hypothesize that NOX plays a role in the regulation of cnidarian–Symbiodiniaceae symbiosis and the host phagosomal maturation process. We investigated NOX function in relation to symbiotic state and heat stress in the sea anemone Exaiptasia diaphana (commonly called aiptasia), a model for cnidarian–Symbiodiniaceae symbiosis and dysbiosis. Our findings show that NOX gene and protein expression is suppressed in the symbiotic state, supporting the hypothesis that symbionts modulate host innate immunity. However, upon heat treatment, we observed increased NOX expression and activity along with NOX localization around algal symbionts, suggesting that host phagosomal maturation processes are engaged during bleaching. We propose a model where the phagocytic NOX complex becomes activated during symbiosis breakdown and bleaching. Our findings support the hypothesis that in situ degradation, facilitated by ROS generated by NOX, plays a key role in the process of dysbiosis. This work contributes to our understanding of cnidarian innate immunity, highlighting critical steps in dysbiosis and phagosomal maturation processes within cnidarian–Symbiodiniaceae symbiosis.

Many cnidarians rely on their dinoflagellate partners from the family Symbiodiniaceae for their ecological success. Symbiotic species of Symbiodiniaceae have two distinct life stages: inside the host, in hospite , and outside the host, ex hospite . Several aspects of cnidarian-algal symbiosis can be understood by comparing these two life stages. Most commonly, algae in culture are used in comparative studies to represent the ex hospite life stage, however, nutrition becomes a confounding variable for this comparison because algal culture media is nutrient rich, while algae in hospite are sampled from hosts maintained in oligotrophic seawater. In contrast to cultured algae, expelled algae may be a more robust representation of the ex hospite state, as the host and expelled algae are in the same seawater environment, removing differences in culture media as a confounding variable. Here, we studied the physiology of algae released from the sea anemone Exaiptasia diaphana (commonly called Aiptasia), a model system for the study of coral-algal symbiosis. In Aiptasia, algae are released in distinct pellets, referred to as egesta, and we explored its potential as an experimental system to represent Symbiodiniaceae in the ex hospite state. Observation under confocal and differential interference contrast microscopy revealed that egesta contained discharged nematocysts, host tissue, and were populated by a diversity of microbes, including protists and cyanobacteria. Further experiments revealed that egesta were released at night. In addition, algae in egesta had a higher mitotic index than algae in hospite , were photosynthetically viable for at least 48 hrs after expulsion, and could competently establish symbiosis with aposymbiotic Aiptasia. We then studied the gene expression of nutrient-related genes and studied their expression using qPCR. From the genes tested, we found that algae from egesta closely mirrored gene expression profiles of algae in hospite and were dissimilar to those of cultured algae, suggesting that algae from egesta are in a nutritional environment that is similar to their in hospite counterparts. Altogether, evidence is provided that algae from Aiptasia egesta are a robust representation of Symbiodiniaceae in the ex hospite state and their use in experiments can improve our understanding of cnidarian-algal symbiosis.

Intracellular photosymbiosis has evolved across life and forms the foundation of coral reef ecosystems. Using the sea anemone Aiptasia as a model, we generated a high-quality proteome of the symbiosome, the organelle that houses algal symbionts. This proteome revealed protein trafficking mechanisms and the types of biomolecules exchanged during symbiosis. Symbiosomal enrichment of lysosomal proteins, visualization of lysosomal fusion, along with reduced symbiosis following knockdown of lysosomal genes, supports its phagolysosomal identity and that extensive co-option of lysosomal proteins shapes the symbiosome. CRISPR/Cas9-induced mutations in the symbiosomal and lysosomal bicarbonate/sulfate transporter, SLC26A11, disrupted symbiosis in both Aiptasia and a reef-building coral. These findings support that anemones and corals independently evolved a carbon-concentrating and sulfate transport mechanism to fuel photosymbiosis by co-opting an orthologous lysosomal transporter.

Identification of driver genes contributes to the understanding of cancer etiology and is imperative for the development of individualized therapies. Gene amplification is a major event in oncogenesis. Driver genes with tumor-specific amplification-dependent overexpression can be therapeutic targets. In this study, we aimed to identify amplification-dependent driver genes in 1,454 solid tumors, across more than 15 cancer types, by integrative analysis of gene expression and copy number. Amplification-dependent overexpression of 64 known driver oncogenes were found in 587 tumors (40%); genes frequently observed were MYC (25%) and MET (18%) in colorectal cancer; SKP2 (21%) in lung squamous cell carcinoma; HIST1H3B (19%) and MYCN (13%) in liver cancer; KIT (57%) in gastrointestinal stromal tumors; and FOXL2 (12%) in squamous cell carcinoma across tissues. Genomic aberrations in 138 known cancer driver genes and 491 established fusion genes were found in 1,127 tumors (78%). Further analyses of 820 cancer-related genes revealed 16 as potential driver genes, with amplification-dependent overexpression restricted to the remaining 22% of samples (327 tumors) initially undetermined genetic drivers. Among them, AXL , which encodes a receptor tyrosine kinase, was recurrently overexpressed and amplified in sarcomas. Our studies of amplification-dependent overexpression identified potential drug targets in individual tumors.

This study aimed to establish the Japanese Cancer Genome Atlas (JCGA) using data from fresh frozen tumor tissues obtained from 5143 Japanese cancer patients, including those with colorectal cancer (31.6%), lung cancer (16.5%), gastric cancer (10.8%) and other cancers (41.1%). The results are part of a single‐center study called “High‐tech Omics‐based Patient Evaluation” or “Project HOPE” conducted at the Shizuoka Cancer Center, Japan. All DNA samples and most RNA samples were analyzed using whole‐exome sequencing, cancer gene panel sequencing, fusion gene panel sequencing and microarray gene expression profiling, and the results were annotated using an analysis pipeline termed “Shizuoka Multi‐omics Analysis Protocol” developed in‐house. Somatic driver alterations were identified in 72.2% of samples in 362 genes (average, 2.3 driver events per sample). Actionable information on drugs that is applicable in the current clinical setting was associated with 11.3% of samples. When including those drugs that are used for investigative purposes, actionable information was assigned to 55.0% of samples. Germline analysis revealed pathogenic mutations in hereditary cancer genes in 9.2% of samples, among which 12.2% were confirmed as pathogenic mutations by confirmatory test. Pathogenic mutations associated with non–cancerous hereditary diseases were detected in 0.4% of samples. Tumor mutation burden (TMB) analysis revealed 5.4% of samples as having the hypermutator phenotype (TMB ≥ 20). Clonal hematopoiesis was observed in 8.4% of samples. Thus, the JCGA dataset and the analytical procedures constitute a fundamental resource for genomic medicine for Japanese cancer patients. The present study aims to establish the Japanese Cancer Genome Atlas (JCGA) by analyzing fresh frozen tumor tissues obtained from 5143 Japanese cancer patients. Somatic driver and druggable alterations were detected in 72.2% and 11.3% of samples, respectively, and germline pathogenic mutations in hereditary cancer genes were identified in 9.2% of samples. The JCGA dataset and analytical procedures constitute a fundamental resource for genomic medicine for Japanese cancer patients.

Whole‐genome analyses have revealed that large‐scale structural variations (SVs) such as whole‐genome duplication (WGD) occur early in the development of many cancers. However, the diversity of chromosomal abnormalities within tumors before and after WGD remains poorly understood. Here, we analyzed various types of Japanese tumor genomes via whole‐genome sequencing and examined the diversity of WGD by focusing on large SVs at the chromosomal level. WGD was detected in 52% of cases, while the frequency of chromothripsis (CT) was 20%. Although aneuploidy via deletion of chromosome arms was common in many cancers, in rare ovarian cancers, all chromosomes were near‐haploidy before WGD. Minor allele analysis revealed that many non‐mutated ohnolog genes drifted down chromosome arms after WGD and returned to normal ploidy, but only 17p, including TP53, which is also an ohnolog, underwent loss of heterozygosity due to arm deletion before WGD in most cancers. TP53 mutations were frequently detected in WGD and CT‐positive tumors, and these SVs strongly correlated with homologous recombination deficiency scores. Furthermore, these tumors had many mutations that continued to generate neoantigens and resulted in worse survival outcomes. Diversity analysis of tumors with WGD will provide a new perspective on structural abnormalities in tumor genomes. Japanese tumor genomes with whole‐genome duplication are diversified by allele imbalance and chromothripsis. These structural abnormalities are linked to a poor prognosis for cancer patients.

Nicotine is metabolized to cotinine by CYP2A6. CYP2A6 genotypes are associated with variations in enzymatic activity and increase the risk of developing tobacco-related diseases, including lung cancer. The high sequence similarity between CYP2A6 and CYP2A7 , combined with frequent duplications, deletions, and structural variants (SVs), such as hybrids, complicate the accurate determination of CYP2A7/CYP2A6 sequence. In this study, we investigated CYP2A6 SVs in 20 Japanese patients with solid cancers using long-read whole-genome sequencing (lrWGS) and short-read whole-genome sequencing (srWGS). Combined lrWGS and srWGS facilitated the detection of copy number variations and SVs, including CYP2A7/CYP2A6 hybrids. Three novel CYP2A7/CYP2A6 hybrids were identified and validated via Sanger sequencing. Patients with CYP2A6 deletion exhibited a lower tumor mutation burden (TMB) than those with normal CYP2A6 . There was no increase in TMB in tumors other than lung cancer in smokers, even when germline CYP2A6 was retained, indicating that the identified germline hybrids affect mutation accumulation in lung cancer in patients with a history of smoking. Overall, combined srWGS-lrWGS facilitated the precise determination of SVs between CYP2A6 and CYP2A7 . This approach could contribute to SV identification in highly homologous chromosomal regions and elucidation of functions of germline SVs in cytochrome P450 in lung cancer.

Despite the frequent detection of KRAS driver mutations in patients with colorectal cancer (CRC), no effective treatments that target mutant KRAS proteins have been introduced into clinical practice. In this study, we identified potential effector molecules, based on differences in gene expression between CRC patients carrying wild-type KRAS (n = 390) and those carrying KRAS mutations in codon 12 (n = 240). CRC patients with wild-type KRAS harboring mutations in HRAS, NRAS, PIK3CA, PIK3CD, PIK3CG, RALGDS, BRAF, or ARAF were excluded from the analysis. At least 11 promising candidate molecules showed greater than two-fold change between the KRAS G12 mutant and wild-type and had a Benjamini-Hochberg-adjusted P value of less than 1E-08, evidence of significantly differential expression between these two groups. Among these 11 genes examined in cell lines transfected with KRAS G12 mutants, BMP4, PHLDA1, and GJB5 showed significantly higher expression level in KRAS G12A, G12D, and G12V transfected cells than in the wild-type transfected cells. We expect that this study will lead to the development of novel treatments that target signaling molecules functioning with KRAS G12-driven CRC.

Tumor mutational burden (TMB) and mutational signatures reflect the process of mutation accumulation in cancer. However, the significance of these emerging characteristics remains unclear. In the present study, we used whole‐exome sequencing to analyze the TMB and mutational signature in solid tumors of 4046 Japanese patients. Eight predominant signatures—microsatellite instability, smoking, POLE, APOBEC, UV, mismatch repair, double‐strand break repair, and Signature 16—were observed in tumors with TMB higher than 1.0 mutation/Mb, whereas POLE and UV signatures only showed moderate correlation with TMB, suggesting the extensive accumulation of mutations due to defective POLE and UV exposure. The contribution ratio of Signature 16, which is associated with hepatocellular carcinoma in drinkers, was increased in hypopharynx cancer. Tumors with predominant microsatellite instability signature were potential candidates for treatment with immune checkpoint inhibitors such as pembrolizumab and were found in 2.8% of cases. Furthermore, based on microarray analysis, tumors with predominant signatures were classified into 2 subgroups depending on the expression of immune‐related genes reflecting differences in the immune context of the tumor microenvironment. Tumor subpopulations differing in the content of infiltrating immune cells might respond differently to immunotherapeutics. An understanding of cancer characteristics based on TMB and mutational signatures could provide new insights into mutation‐driven tumorigenesis. By applying whole‐exome sequencing and microarray technologies for high‐throughput genomic and transcriptional analysis, this study investigates the mutational burden and signatures in a large cohort of tumor samples of different histology from Japanese patients. The contribution ratio of Signature 16, which is associated with hepatocellular carcinoma in drinkers, was increased in hypopharynx cancer. Furthermore, based on microarray analysis, tumors with predominant signatures were classified into 2 subgroups depending on the expression of immune‐related genes, reflecting differences in the immune context of the tumor microenvironment.

Tumor mutational burden analysis using whole‐exome sequencing highlights features of tumors with various mutations or known driver alterations. Cancers with few changes in the exon regions have unclear characteristics, even though low‐mutated tumors are often detected in pan‐cancer analysis. In the present study, we analyzed tumors with low tumor mutational burden listed in the Japanese version of The Cancer Genome Atlas, a data set of 5020 primary solid tumors. Our analysis revealed that detection rates of known driver mutations and copy number variation were decreased in samples with tumor mutational burden below 1.0 (ultralow tumor), compared with those in samples with low tumor mutational burden (≤5 mutations/Mb). This trend was also observed in The Cancer Genome Atlas data set. In the ultralow tumor mutational burden tumors, expression analysis showed decreased TP53 inactivation and chromosomal instability. TP53 inactivation frequently correlated with PI3K/mTOR‐related gene expression, implying suppression of the PI3K/mTOR pathway in ultralow tumor mutational burden tumors. In common with mutational burden, the T cell‐inflamed gene expression profiling signature was a potential marker for prediction of an immune checkpoint inhibitor response, and some ultralow tumor mutational burden tumor populations highly expressed this signature. Our analysis focused on how these tumors could provide insight into tumors with low somatic alteration that are difficult to detect solely using whole‐exome sequencing. This study describes our analysis of tumors with a low tumor burden level in the Japanese version of The Cancer Genome Atlas (JCGA), a data set of 5020 primary solid tumors. In the ultralow tumor mutational burden tumors, expression analysis showed decreased TP53 inactivation and chromosomal instability. Our analysis focuses on how these tumors can provide insight into tumors with low somatic alteration that are difficult to detect solely by whole exome sequencing.