SUMMARY Lineage mapping has identified both proliferative and quiescent intestinal stem cells, but the molecular circuitry controlling stem cell quiescence is incompletely understood. By lineage mapping, we show Lrig1, a pan-ErbB inhibitor, marks predominately non-cycling, long-lived stem cells located at the crypt base that, upon injury, proliferate and divide to replenish damaged crypts. Transcriptome profiling of Lrig1+ colonic stem cells differs markedly from highly proliferative, Lgr5+ colonic stem cells; genes up-regulated in the Lrig1+ population include those involved in cell cycle repression and response to oxidative damage. Loss of Apc in Lrig1+ cells leads to intestinal adenomas and genetic ablation of Lrig1 results in heightened ErbB1-3 expression and duodenal adenomas. These results shed light on the relationship between proliferative and quiescent intestinal stem cells, and support a model in which intestinal stem cell quiescence is maintained by calibrated ErbB signaling with loss of a negative regulator predisposing to neoplasia.

Mutant KRAS colorectal cancer (CRC) cells release protein-laden exosomes that can alter the tumor microenvironment. To test whether exosomal RNAs also contribute to changes in gene expression in recipient cells, and whether mutant KRAS might regulate the composition of secreted microRNAs (miRNAs), we compared small RNAs of cells and matched exosomes from isogenic CRC cell lines differing only in KRAS status. We show that exosomal profiles are distinct from cellular profiles, and mutant exosomes cluster separately from wild-type KRAS exosomes. miR-10b was selectively increased in wild-type exosomes, while miR-100 was increased in mutant exosomes. Neutral sphingomyelinase inhibition caused accumulation of miR-100 only in mutant cells, suggesting KRAS-dependent miRNA export. In Transwell co-culture experiments, mutant donor cells conferred miR-100-mediated target repression in wild-type-recipient cells. These findings suggest that extracellular miRNAs can function in target cells and uncover a potential new mode of action for mutant KRAS in CRC.DOI: http://dx.doi.org/10.7554/eLife.07197.001

Summary Decreasing glucagon action lowers the blood glucose and may be useful therapeutically for diabetes. However, interrupted glucagon signaling leads to α-cell proliferation. To identify postulated hepatic-derived, circulating factor(s) responsible for α-cell proliferation, we used transcriptomics/proteomics/metabolomics in three models of interrupted glucagon signaling and found that proliferation of mouse, zebrafish, and human α-cells was mTOR- and FoxP transcription factor-dependent. Changes in hepatic amino acid (AA) catabolism gene expression predicted the observed increase in circulating AA. Mimicking these AA levels stimulated α-cell proliferation in a newly developed in vitro assay with L-glutamine being a critical AA. α-cell expression of the AA transporter Slc38a5 was markedly increased in mice with interrupted glucagon signaling and played a role in α-cell proliferation. These results indicate a hepatic-α-islet cell axis where glucagon regulates serum AA availability and AA, especially L-glutamine, regulates α-cell proliferation and mass via mTOR-dependent nutrient sensing.

Recent studies have shown that circular RNAs (circRNAs) are abundant, widely expressed in mammals, and can display cell-type specific expression. However, how production of circRNAs is regulated and their precise biological function remains largely unknown. To study how circRNAs might be regulated during colorectal cancer progression, we used three isogenic colon cancer cell lines that differ only in KRAS mutation status. Cellular RNAs from the parental DLD-1 cells that contain both wild-type and G13D mutant KRAS alleles and isogenically-matched derivative cell lines, DKO-1 (mutant KRAS allele only) and DKs-8 (wild-type KRAS allele only) were analyzed using RNA-Seq. We developed a bioinformatics pipeline to identify and evaluate circRNA candidates from RNA-Seq data. Hundreds of high-quality circRNA candidates were identified in each cell line. Remarkably, circRNAs were significantly down-regulated at a global level in DLD-1 and DKO-1 cells compared to DKs-8 cells, indicating a widespread effect of mutant KRAS on circRNA abundance. This finding was confirmed in two independent colon cancer cell lines HCT116 (KRAS mutant) and HKe3 (KRAS WT). In all three cell lines, circRNAs were also found in secreted extracellular-vesicles, and circRNAs were more abundant in exosomes than cells. Our results suggest that circRNAs may serve as promising cancer biomarkers.

Many patients with type 1 diabetes (T1D) have residual β cells producing small amounts of C-peptide long after disease onset but develop an inadequate glucagon response to hypoglycemia following T1D diagnosis. The features of these residual β cells and α cells in the islet endocrine compartment are largely unknown, due to the difficulty of comprehensive investigation. By studying the T1D pancreas and isolated islets, we show that remnant β cells appeared to maintain several aspects of regulated insulin secretion. However, the function of T1D α cells was markedly reduced, and these cells had alterations in transcription factors constituting α and β cell identity. In the native pancreas and after placing the T1D islets into a non-autoimmune, normoglycemic in vivo environment, there was no evidence of α-to-β cell conversion. These results suggest an explanation for the disordered T1D counterregulatory glucagon response to hypoglycemia.