Transcriptome Analysis during Human Trophectoderm Specification Suggests New Roles of Metabolic and Epigenetic Genes

In humans, successful pregnancy depends on a cascade of dynamic events during early embryonic development. Unfortunately, molecular data on these critical events is scarce. To improve our understanding of the molecular mechanisms that govern the specification/development of the trophoblast cell lineage, the transcriptome of human trophectoderm (TE) cells from day 5 blastocysts was compared to that of single day 3 embryos from our in vitro fertilization program by using Human Genome U133 Plus 2.0 microarrays. Some of the microarray data were validated by quantitative RT-PCR. The TE molecular signature included 2,196 transcripts, among which were genes already known to be TE-specific (GATA2, GATA3 and GCM1) but also genes involved in trophoblast invasion (MUC15), chromatin remodeling (specifically the DNA methyltransferase DNMT3L) and steroid metabolism (HSD3B1, HSD17B1 and FDX1). In day 3 human embryos 1,714 transcripts were specifically up-regulated. Besides stemness genes such as NANOG and DPPA2, this signature included genes belonging to the NLR family (NALP4, 5, 9, 11 and 13), Ret finger protein-like family (RFPL1, 2 and 3), Melanoma Antigen family (MAGEA1, 2, 3, 5, 6 and 12) and previously unreported transcripts, such as MBD3L2 and ZSCAN4. This study provides a comprehensive outlook of the genes that are expressed during the initial embryo-trophectoderm transition in humans. Further understanding of the biological functions of the key genes involved in steroidogenesis and epigenetic regulation of transcription that are up-regulated in TE cells may clarify their contribution to TE specification and might also provide new biomarkers for the selection of viable and competent blastocysts

Regulation of CD44 expression by bone morphogenetic protein (BMP) signaling and inflammatory stimuli

The adhesion molecule CD44 is involved in inflammation-mediated gastric carcinogenesis. Lgr5 marks populations of gastric stem cells. Lgr5-expressing cells can be identified in the antrum, along the lesser curvature of the oxyntic mucosa, and in gastric carcinomas. We reported that inflammation and inhibition of BMP signaling, achieved by expression of the BMP inhibitor noggin (Nog) in the oxyntic epithelium (H+/K+-Nog mice), and by crossing Lgr5-Cre mice to mice with floxed alleles of BMP receptor 1A (Lgr5-Cre;BMPR1Aflox-flox mice), activate aberrant Lgr5 cells that give rise to dysplastic, proliferating lineages. We tested the hypothesis that BMP signaling regulates the fate of Lgr5 cells and the expression of CD44 during gastric inflammation and carcinogenesis. To conduct lineage tracing in the presence of inhibition of BMP signaling we crossed H+/K+-Nog mice and BMPR1Aflox-flox mice to Rosa26-tdTomato (Tom) reporter mice to generate H+/K+-Nog;Rosa26-tdTom and BMPR1Aflox-flox;Rosa26-tdTom mice. Both mice were crossed to Lgr5-Cre mice to generate Lgr5-Cre;H+/K+-Nog;Rosa26-tdTom and Lgr5-Cre;BMPR1Aflox-flox;Rosa26tdTom mice. Infection with H. felis (HF) was used to induce inflammation. Animals were analyzed 3 months after HF inoculation. Distribution of Tom-, CD44- and BrdU-positive cells was analyzed by immunostaining. Cultures of human organoids derived from antral and fundic biopsies were used to assess the effect of IFNγ (30 ng/ml) on the expression of CD44 and Lgr5. Expression of CD44, Lgr5, IFNγ, TNF-α, IL-1-β, and of the BMP signaling inhibitors Smad6, Smad7 and Bambi was measured in 16 human tumors of the lesser curvature and in an equal number of cancers of the body using QRT-PCR. Inhibition of BMP signaling and infection with HF induced the expression of CD44. HF-infected Lgr5-Cre;H+/K+-Nog;Rosa26-tdTom mice and Lgr5-Cre;BMPR1Aflox-flox;Rosa26-tdTom mice exhibited Tom expression in glands of the antrum and of the lesser curvature that positively stained with anti-CD44 and -BrdU antibodies. IFNγ induced the expression of CD44 and Lgr5 in antral, but not in fundic organoids. Expression of CD44, Lgr5, IFNγ, TNF-α, IL-1-β, Smad6, Smad7 and Bambi was increased in tumors of the lesser curvature but not of the body. Inflammation and inhibition of BMP signaling induce CD44 and Lgr5 in vivo and in vitro, in both mouse and human tissues. Lineage tracing studies demonstrate the presence of aberrant Lgr5 cells located along the lesser curvature that give rise to proliferating lineages that express CD44. Tumors of the lesser curvature but not of the body exhibit increased expression of CD44, Lgr5, BMP inhibitors, and pro-inflammatory molecules. These findings underscore the importance of inflammation and BMP signaling in the development of neoplastic lesions of the stomach

Less is more: Nutrient limitation induces cross-talk of nutrient sensing pathways with NAD+ homeostasis and contributes to longevity

Nutrient sensing pathways and their regulation grant cells control over their metabolism and growth in response to changing nutrients. Factors that regulate nutrient sensing can also modulate longevity. Reduced activity of nutrient sensing pathways such as glucose-sensing PKA, nitrogen-sensing TOR and S6 kinase homolog Sch9 have been linked to increased life span in the yeast, Saccharomyces cerevisiae, and higher eukaryotes. Recently, reduced activity of amino acid sensing SPS pathway was also shown to increase yeast life span. Life span extension by reduced SPS activity requires enhanced NAD(+) (nicotinamide adenine dinucleotide, oxidized form) and nicotinamide riboside (NR, a NAD(+) precursor) homeostasis. Maintaining adequate NAD(+) pools has been shown to play key roles in life span extension, but factors regulating NAD(+) metabolism and homeostasis are not completely understood. Recently, NAD(+) metabolism was also linked to the phosphate (Pi)-sensing PHO pathway in yeast. Canonical PHO activation requires Pi-starvation. Interestingly, NAD(+) depletion without Pi-starvation was sufficient to induce PHO activation, increasing NR production and mobilization. Moreover, SPS signaling appears to function in parallel with PHO signaling components to regulate NR/NAD(+) homeostasis. These studies suggest that NAD(+) metabolism is likely controlled by and/or coordinated with multiple nutrient sensing pathways. Indeed, cross-regulation of PHO, PKA, TOR and Sch9 pathways was reported to potentially affect NAD(+) metabolism; though detailed mechanisms remain unclear. This review discusses yeast longevity-related nutrient sensing pathways and possible mechanisms of life span extension, regulation of NAD(+) homeostasis, and cross-talk among nutrient sensing pathways and NAD(+) homeostasis