BackgroundIdentification of genes with invariant levels of gene expression is a prerequisite for validating transcriptomic changes accompanying development. Ideally expression of these genes should be independent of the morphogenetic process or environmental condition tested as well as the methods used for RNA purification and analysis.ResultsIn an effort to identify endogenous genes meeting these criteria nine reference genes (RG) were tested in two Petunia lines (Mitchell and V30). Growth conditions differed in Mitchell and V30, and different methods were used for RNA isolation and analysis. Four different software tools were employed to analyze the data. We merged the four outputs by means of a non-weighted unsupervised rank aggregation method. The genes identified as optimal for transcriptomic analysis of Mitchell and V30 were EF1α in Mitchell and CYP in V30, whereas the least suitable gene was GAPDH in both lines.ConclusionsThe least adequate gene turned out to be GAPDH indicating that it should be rejected as reference gene in Petunia. The absence of correspondence of the best-suited genes suggests that assessing reference gene stability is needed when performing normalization of data from transcriptomic analysis of flower and leaf development.

BackgroundThe Cancer Genome Atlas (TCGA) offers a multilayered view of genomics and epigenomics data of many human cancer types. However, the retrieval of expression and methylation data from TCGA is a cumbersome and time-consuming task.ResultsWanderer is an intuitive Web tool allowing real time access and visualization of gene expression and DNA methylation profiles from TCGA. Given a gene query and selection of a TCGA dataset (e.g., colon adenocarcinomas), the Web resource provides the expression profile, at the single exon level, and the DNA methylation levels of HumanMethylation450 BeadChip loci inside or in the vicinity of the queried gene. Graphic and table outputs include individual and summary data as well as statistical tests, allowing the comparison of tumor and normal profiles and the exploration along the genomic locus and across tumor collections.ConclusionsWanderer offers a simple interface to straightforward access to TCGA data, amenable to experimentalists and clinicians without bioinformatics expertise. Wanderer may be accessed at http://maplab.cat/wanderer.

SUMMARY We addressed the precursor role of aging-like spontaneous promoter DNA hypermethylation in initiating tumorigenesis. Using mouse colon-derived organoids, we show that promoter hypermethylation spontaneously arises in cells mimicking the human aging-like phenotype. The silenced genes activate the Wnt pathway, causing a stem-like state and differentiation defects. These changes render aged organoids profoundly more sensitive than young ones to transformation by BrafV600E, producing the typical human proximal BRAFV600E-driven colon adenocarcinomas characterized by extensive, abnormal gene-promoter CpG-island methylation, or the methylator phenotype (CIMP). Conversely, CRISPR-mediated simultaneous inactivation of a panel of the silenced genes markedly sensitizes to BrafV600E-induced transformation. Our studies tightly link aging-like epigenetic abnormalities to intestinal cell fate changes and predisposition to oncogene-driven colon tumorigenesis.

Neural stem cells (NSCs) generate neurons throughout life in the mammalian hippocampus. However, the potential for long-term self-renewal of individual NSCs within the adult brain remains unclear. We used 2-photon microscopy and followed NSCs that were genetically labeled through conditional recombination driven by the regulatory elements of the stem cell-expressed genes GLI Family Zinc Finger 1 (Gli1) or Achaete-scute homolog 1 (Ascl1). Through intravital imaging of NSCs and their progeny, we identify a population of Gli1-targeted NSCs showing long-term self-renewal in the adult hippocampus. In contrast, once activated, Ascl1-targeted NSCs undergo limited proliferative activity before they become exhausted. Using single-cell RNA sequencing, we show that Gli1- and Ascl1-targeted cells have highly similar yet distinct transcriptional profiles, supporting the existence of heterogeneous NSC populations with diverse behavioral properties. Thus, we here identify long-term self-renewing NSCs that contribute to the generation of new neurons in the adult hippocampus.