Biochemical properties of chromatin domains define genome compartmentalization.

Chromatin three-dimensional (3D) organization inside the cell nucleus determines the separation of euchromatin and heterochromatin domains. Their segregation results in the definition of active and inactive chromatin compartments, whereby the local concentration of associated proteins, RNA and DNA results in the formation of distinct subnuclear structures. Thus, chromatin domains spatially confined in a specific 3D nuclear compartment are expected to share similar epigenetic features and biochemical properties, in terms of accessibility and solubility. Based on this rationale, we developed the 4f-SAMMY-seq to map euchromatin and heterochromatin based on their accessibility and solubility, starting from as little as 10 000 cells. Adopting a tailored bioinformatic data analysis approach we reconstruct also their 3D segregation in active and inactive chromatin compartments and sub-compartments, thus recapitulating the characteristic properties of distinct chromatin states. A key novelty of the new method is the capability to map both the linear segmentation of open and closed chromatin domains, as well as their compartmentalization in one single experiment.

A PET-Surrogate Signature for the Interrogation of the Metabolic Status of Breast Cancers.

Metabolic alterations in cancers can be exploited for diagnostic, prognostic, and therapeutic purposes. This is exemplified by 18F-fluorodeoxyglucose (FDG)-positron emission tomography (FDG-PET), an imaging tool that relies on enhanced glucose uptake by tumors for diagnosis and staging. By performing transcriptomic analysis of breast cancer (BC) samples from patients stratified by FDG-PET, a 54-gene signature (PETsign) is identified that recapitulates FDG uptake. PETsign is independently prognostic of clinical outcome in luminal BCs, the most common and heterogeneous BC molecular subtype, which requires improved stratification criteria to guide therapeutic decision-making. The prognostic power of PETsign is stable across independent BC cohorts and disease stages including the earliest BC stage, arguing that PETsign is an ab initio metabolic signature. Transcriptomic and metabolomic analysis of BC cells reveals that PETsign predicts enhanced glycolytic dependence and reduced reliance on fatty acid oxidation. Moreover, coamplification of PETsign genes occurs frequently in BC arguing for their causal role in pathogenesis. CXCL8 and EGFR signaling pathways feature strongly in PETsign, and their activation in BC cells causes a shift toward a glycolytic phenotype. Thus, PETsign serves as a molecular surrogate for FDG-PET that could inform clinical management strategies for BC patients.

RAGE engagement by SARS-CoV-2 enables monocyte infection and underlies COVID-19 severity.

The spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has fueled the COVID-19 pandemic with its enduring medical and socioeconomic challenges because of subsequent waves and long-term consequences of great concern. Here, we chart the molecular basis of COVID-19 pathogenesis by analyzing patients' immune responses at single-cell resolution across disease course and severity. This approach confirms cell subpopulation-specific dysregulation in COVID-19 across disease course and severity and identifies a severity-associated activation of the receptor for advanced glycation endproducts (RAGE) pathway in monocytes. In vitro THP1-based experiments indicate that monocytes bind the SARS-CoV-2 S1-receptor binding domain (RBD) via RAGE, pointing to RAGE-Spike interaction enabling monocyte infection. Thus, our results demonstrate that RAGE is a functional receptor of SARS-CoV-2 contributing to COVID-19 severity.

The genomic landscape of canine diffuse large B-cell lymphoma identifies distinct subtypes with clinical and therapeutic implications.

Diffuse large B-cell lymphoma (DLBCL) is the most common lymphoid neoplasm in dogs and in humans. It is characterized by a remarkable degree of clinical heterogeneity that is not completely elucidated by molecular data. This poses a major barrier to understanding the disease and its response to therapy, or when treating dogs with DLBCL within clinical trials. We performed an integrated analysis of exome (n = 77) and RNA sequencing (n = 43) data in a cohort of canine DLBCL to define the genetic landscape of this tumor. A wide range of signaling pathways and cellular processes were found in common with human DLBCL, but the frequencies of the most recurrently mutated genes (TRAF3, SETD2, POT1, TP53, MYC, FBXW7, DDX3X and TBL1XR1) differed. We developed a prognostic model integrating exonic variants and clinical and transcriptomic features to predict the outcome in dogs with DLBCL. These results comprehensively define the genetic drivers of canine DLBCL and can be prospectively utilized to identify new therapeutic opportunities.

GZMKhigh CD8+ T effector memory cells are associated with CD15high neutrophil abundance in non-metastatic colorectal tumors and predict poor clinical outcome.

CD8+ T cells are a major prognostic determinant in solid tumors, including colorectal cancer (CRC). However, understanding how the interplay between different immune cells impacts on clinical outcome is still in its infancy. Here, we describe that the interaction of tumor infiltrating neutrophils expressing high levels of CD15 with CD8+ T effector memory cells (TEM) correlates with tumor progression. Mechanistically, stromal cell-derived factor-1 (CXCL12/SDF-1) promotes the retention of neutrophils within tumors, increasing the crosstalk with CD8+ T cells. As a consequence of the contact-mediated interaction with neutrophils, CD8+ T cells are skewed to produce high levels of GZMK, which in turn decreases E-cadherin on the intestinal epithelium and favors tumor progression. Overall, our results highlight the emergence of GZMKhigh CD8+ TEM in non-metastatic CRC tumors as a hallmark driven by the interaction with neutrophils, which could implement current patient stratification and be targeted by novel therapeutics.

Gut Microbiota Functional Dysbiosis Relates to Individual Diet in Subclinical Carotid Atherosclerosis.

Gut Microbiota (GM) dysbiosis associates with Atherosclerotic Cardiovascular Diseases (ACVD), but whether this also holds true in subjects without clinically manifest ACVD represents a challenge of personalized prevention. We connected exposure to diet (self-reported by food diaries) and markers of Subclinical Carotid Atherosclerosis (SCA) with individual taxonomic and functional GM profiles (from fecal metagenomic DNA) of 345 subjects without previous clinically manifest ACVD. Subjects without SCA reported consuming higher amounts of cereals, starchy vegetables, milky products, yoghurts and bakery products versus those with SCA (who reported to consume more mechanically separated meats). The variety of dietary sources significantly overlapped with the separations in GM composition between subjects without SCA and those with SCA (RV coefficient between nutrients quantities and microbial relative abundances at genus level = 0.65, p-value = 0.047). Additionally, specific bacterial species (Faecalibacterium prausnitzii in the absence of SCA and Escherichia coli in the presence of SCA) are directly related to over-representation of metagenomic pathways linked to different dietary sources (sulfur oxidation and starch degradation in absence of SCA, and metabolism of amino acids, syntheses of palmitate, choline, carnitines and Trimethylamine n-oxide in presence of SCA). These findings might contribute to hypothesize future strategies of personalized dietary intervention for primary CVD prevention setting.

Identification of MYC-Dependent Transcriptional Programs in Oncogene-Addicted Liver Tumors.

Tumors driven by activation of the transcription factor MYC generally show oncogene addiction. However, the gene expression programs that depend upon sustained MYC activity remain unknown. In this study, we employed a mouse model of liver carcinoma driven by a reversible tet-MYC transgene, combined with chromatin immunoprecipitation and gene expression profiling to identify MYC-dependent regulatory events. As previously reported, MYC-expressing mice exhibited hepatoblastoma- and hepatocellular carcinoma-like tumors, which regressed when MYC expression was suppressed. We further show that cellular transformation, and thus initiation of liver tumorigenesis, were impaired in mice harboring a MYC mutant unable to associate with the corepressor protein MIZ1 (ZBTB17). Notably, switching off the oncogene in advanced carcinomas revealed that MYC was required for the continuous activation and repression of distinct sets of genes, constituting no more than half of all genes deregulated during tumor progression and an even smaller subset of all MYC-bound genes. Altogether, our data provide the first detailed analysis of a MYC-dependent transcriptional program in a fully developed carcinoma and offer a guide to identifying the critical effectors contributing to MYC-driven tumor maintenance. Cancer Res; 76(12); 3463-72. ©2016 AACR.

Genome-wide analysis of p53 transcriptional programs in B cells upon exposure to genotoxic stress in vivo.

The tumor suppressor p53 is a transcription factor that coordinates the cellular response to DNA damage. Here we provide an integrated analysis of p53 genomic occupancy and p53-dependent gene regulation in the splenic B and non-B cell compartments of mice exposed to whole-body ionizing radiation, providing insight into general principles of p53 activity in vivo. In unstressed conditions, p53 bound few genomic targets; induction of p53 by ionizing radiation increased the number of p53 bound sites, leading to highly overlapping profiles in the different cell types. Comparison of these profiles with chromatin features in unstressed B cells revealed that, upon activation, p53 localized at active promoters, distal enhancers, and a smaller set of unmarked distal regions. At promoters, recognition of the canonical p53 motif as well as binding strength were associated with p53-dependent transcriptional activation, but not repression, indicating that the latter was most likely indirect. p53-activated targets constituted the core of a cell type-independent response, superimposed onto a cell type-specific program. Core response genes included most of the known p53-regulated genes, as well as many new ones. Our data represent a unique characterization of the p53-regulated response to ionizing radiation in vivo.

Whole exome sequencing identifies driver mutations in asymptomatic computed tomography-detected lung cancers with normal karyotype.

The efficacy of curative surgery for lung cancer could be largely improved by non-invasive screening programs, which can detect the disease at early stages. We previously showed that 18% of screening-identified lung cancers demonstrate a normal karyotype and, following high-density genome scanning, can be subdivided into samples with 1) numerous; 2) none; and 3) few copy number alterations. Whole exome sequencing was applied to the two normal karyotype, screening-detected lung cancers, constituting group 2, as well as normal controls. We identified mutations in both tumors, including KEAP1 (commonly mutated in lung cancers) in one, and TP53, PMS1, and MSH3 (well-characterized DNA-repair genes) in the other. The two normal karyotype screening-detected lung tumors displayed a typical lung cancer mutational profile that only next generation sequencing could reveal, which offered an additional contribution to the over-diagnosis bias concept hypothesized within lung cancer screening programs.