Mutant APC reshapes Wnt signaling plasma membrane nanodomains by altering cholesterol levels via oncogenic ß-catenin.

Although the role of the Wnt pathway in colon carcinogenesis has been described previously, it has been recently demonstrated that Wnt signaling originates from highly dynamic nano-assemblies at the plasma membrane. However, little is known regarding the role of oncogenic APC in reshaping Wnt nanodomains. This is noteworthy, because oncogenic APC does not act autonomously and requires activation of Wnt effectors upstream of APC to drive aberrant Wnt signaling. Here, we demonstrate the role of oncogenic APC in increasing plasma membrane free cholesterol and rigidity, thereby modulating Wnt signaling hubs. This results in an overactivation of Wnt signaling in the colon. Finally, using the Drosophila sterol auxotroph model, we demonstrate the unique ability of exogenous free cholesterol to disrupt plasma membrane homeostasis and drive Wnt signaling in a wildtype APC background. Collectively, these findings provide a link between oncogenic APC, loss of plasma membrane homeostasis and CRC development.

Personalized Nutrition Using Microbial Metabolite Phenotype to Stratify Participants and Non-Invasive Host Exfoliomics Reveal the Effects of Flaxseed Lignan Supplementation in a Placebo-Controlled Crossover Trial.

High-fiber plant foods contain lignans that are converted to bioactive enterolignans, enterolactone (ENL) and enterodiol (END) by gut bacteria. Previously, we conducted an intervention study to gain mechanistic insight into the potential chemoprotective effects of flaxseed lignan supplementation (secoisolariciresinol diglucoside; SDG) compared to a placebo in 42 men and women. Here, we expand on these analyses to further probe the impact of the microbial metabolite phenotype on host gene expression in response to lignan exposure. We defined metabolic phenotypes as high- or low-ENL excretion based on the microbial metabolism of SDG. RNA-seq was used to assess host gene expression in fecal exfoliated cells. Stratified by microbial ENL excretion, differentially expressed (DE) genes in high- and low-ENL excreter groups were compared. Linear discriminant analysis using the ENL phenotypes identified putative biomarker combinations of genes capable of discriminating the lignan treatment from the placebo. Following lignan intervention, a total of 165 DE genes in high-ENL excreters and 1450 DE genes in low-ENL excreters were detected. Functional analysis identified four common upstream regulators (master genes): CD3, IFNG, IGF1 and TNFRSF1A. Our findings suggest that the enhanced conversion of flaxseed lignan to ENL is associated with a suppressed inflammatory status.

Transcriptomic Profiling of Gene Expression Associated with Granulosa Cell Tumor Development in a Mouse Model.

Ovarian granulosa cell tumors (GCTs) are rare sex cord-stromal tumors, accounting for ~5% ovarian tumors. The etiology of GCTs remains poorly defined. Genetically engineered mouse models are potentially valuable for understanding the pathogenesis of GCTs. Mice harboring constitutively active TGFß signaling (TGFBR1-CA) develop ovarian GCTs that phenocopy several hormonal and molecular characteristics of human GCTs. To determine molecular alterations in the ovary upon TGFß signaling activation, we performed transcriptomic profiling of gene expression associated with GCT development using ovaries from 1-month-old TGFBR1-CA mice and age-matched controls. RNA-sequencing and bioinformatics analysis coupled with the validation of select target genes revealed dysregulations of multiple cellular events and signaling molecules/pathways. The differentially expressed genes are enriched not only for known GCT-related pathways and tumorigenic events but also for signaling events potentially mediated by neuroactive ligand-receptor interaction, relaxin signaling, insulin signaling, and complements in TGFBR1-CA ovaries. Additionally, a comparative analysis of our data in mice with genes dysregulated in human GCTs or granulosa cells overexpressing a mutant FOXL2, the genetic hallmark of adult GCTs, identified some common genes altered in both conditions. In summary, this study has revealed the molecular signature of ovarian GCTs in a mouse model that harbors the constitutive activation of TGFBR1. The findings may be further exploited to understand the pathogenesis of a class of poorly defined ovarian tumors.

Single-cell RNA Sequencing Reveals How the Aryl Hydrocarbon Receptor Shapes Cellular Differentiation Potency in the Mouse Colon.

Despite recent progress recognizing the importance of aryl hydrocarbon receptor (Ahr)-dependent signaling in suppressing colon tumorigenesis, its role in regulating colonic crypt homeostasis remains unclear. To assess the effects of Ahr on intestinal epithelial cell heterogeneity and functional phenotypes, we utilized single-cell transcriptomics and advanced analytic strategies to generate a high-quality atlas for colonic intestinal crypts from wild-type and intestinal-specific Ahr knockout mice. Here we observed the promotive effects of Ahr deletion on Foxm1-regulated genes in crypt-associated canonical epithelial cell types and subtypes of goblet cells and deep crypt-secretory cells. We also show that intestinal Ahr deletion elevated single-cell entropy (a measure of differentiation potency or cell stemness) and RNA velocity length (a measure of the rate of cell differentiation) in noncycling and cycling Lgr5+ stem cells. In general, intercellular signaling cross-talk via soluble and membrane-bound factors was perturbed in Ahr-null colonocytes. Taken together, our single-cell RNA sequencing analyses provide new evidence of the molecular function of Ahr in modulating putative stem cell driver genes, cell potency lineage decisions, and cell-cell communication in vivo. PREVENTION RELEVANCE: Our mouse single-cell RNA sequencing analyses provide new evidence of the molecular function of Ahr in modulating colonic stemness and cell-cell communication in vivo. From a cancer prevention perspective, Ahr should be considered a therapeutic target to recalibrate remodeling of the intestinal stem cell niche.

Exfoliated epithelial cell transcriptome reflects both small and large intestinal cell signatures in piglets.

Assessing intestinal development and host-microbe interactions in healthy human infants requires noninvasive approaches. We have shown that the transcriptome of exfoliated epithelial cells in feces can differentiate breast-fed and formula-fed infants and term and preterm infants. However, it is not fully understood which regions of the intestine that the exfoliated cells represent. Herein, the transcriptional profiles of exfoliated cells with that of the ileal and colonic mucosa were compared. We hypothesized that exfoliated cells in the distal colon would reflect mucosal signatures of more proximal regions of the gut. Two-day-old piglets (n = 8) were fed formulas for 20 days. Luminal contents and mucosa were collected from ileum (IL), ascending colon (AC), and descending (DC) colon, and mRNA was extracted and sequenced. On average, ∼13,000 genes were mapped in mucosal tissues and ∼10,000 in luminal contents. The intersection of detected genes between three mucosa regions and DC exfoliome indicated an approximately 99% overlap. On average, 49% of the genes in IL, AC, and DC mucosa were present in the AC and DC exfoliome. Genes expressed predominantly in specific anatomic sites (stomach, pancreas, small intestine, colon) were detectable in exfoliated cells. In addition, gene markers for all intestinal epithelial cell types were expressed in the exfoliome representing a diverse array of cell types arising from both the small and large intestine. Genes were mapped to nutrient absorption and transport and immune function. Thus, the exfoliome represents a robust reservoir of information in which to assess intestinal development and responses to dietary interventions.NEW & NOTEWORTHY The transcriptome of exfoliated epithelial cells in stool contain gene signatures from both small and large intestinal mucosa affording a noninvasive approach to assess gut health and function.