Cycling Lgr5+ stem cells fuel the rapid turnover of the adult intestinal epithelium. The existence of quiescent Lgr5+ cells has been reported, while an alternative quiescent stem cell population is ...believed to reside at crypt position +4. Here, we generated a novel Ki67RFP knock‐in allele that identifies dividing cells. Using Lgr5‐GFP;Ki67RFP mice, we isolated crypt stem and progenitor cells with distinct Wnt signaling levels and cell cycle features and generated their molecular signature using microarrays. Stem cell potential of these populations was further characterized using the intestinal organoid culture. We found that Lgr5high stem cells are continuously in cell cycle, while a fraction of Lgr5low progenitors that reside predominantly at +4 position exit the cell cycle. Unlike fast dividing CBCs, Lgr5low Ki67− cells have lost their ability to initiate organoid cultures, are enriched in secretory differentiation factors, and resemble the Dll1 secretory precursors and the label‐retaining cells of Winton and colleagues. Our findings support the cycling stem cell hypothesis and highlight the cell cycle heterogeneity of early progenitors during lineage commitment.
Synopsis
A Ki67‐RFP knock‐in allele highlights the heterogeneity of Lgr5+ stem‐ and progenitor cells based on in vivo cell cycle profiles.
Ki67‐RFP allele allows identification and visualization of proliferating cells in vivo.
Lgr5high intestinal stem cells are continuously in the cell cycle.
Quiescent Lgr5low “+4” cells are secretory precursors.
A Ki67‐RFP knock‐in allele highlights the heterogeneity of Lgr5+ stem‐ and progenitor cells based on in vivo cell cycle profiles.
The regeneration-capable flatworm
is a powerful model organism to study the biology of stem cells in vivo. As a flatworm amenable to transgenesis, it complements the historically used planarian ...flatworm models, such as
. However, information on the transcriptome and markers of stem cells in
is limited. We generated a de novo transcriptome assembly and performed the first comprehensive characterization of gene expression in the proliferating cells of
, represented by somatic stem cells, called neoblasts, and germline cells. Knockdown of a selected set of neoblast genes, including
,
,
,
, and
, confirmed their crucial role for the functionality of somatic neoblasts during homeostasis and regeneration. The generated
transcriptome assembly and gene expression signatures of somatic neoblasts and germline cells will be a valuable resource for future molecular studies in
.
Colorectal cancer stem cells (CSCs) express Lgr5 and display extensive stem cell-like multipotency and self-renewal and are thought to seed metastatic disease. Here, we used a mouse model of ...colorectal cancer (CRC) and human tumor xenografts to investigate the cell of origin of metastases. We found that most disseminated CRC cells in circulation were Lgr5− and formed distant metastases in which Lgr5+ CSCs appeared. This plasticity occurred independently of stemness-inducing microenvironmental factors and was indispensable for outgrowth, but not establishment, of metastases. Together, these findings show that most colorectal cancer metastases are seeded by Lgr5− cells, which display intrinsic capacity to become CSCs in a niche-independent manner and can restore epithelial hierarchies in metastatic tumors.
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•The majority of disseminating cells of colorectal cancer are Lgr5−•Lgr5− cancer cells are the main seeds of colorectal cancer metastatic lesions•Long-term metastatic growth from Lgr5− cells requires appearance of Lgr5+ cells•Lgr5− metastases have the intrinsic capacity to re-establish the cellular hierarchy
Van Rheenen and colleagues study Lgr5+ cancer stem cells during colorectal cancer metastasis. They demonstrate that the majority of metastases are seeded by Lgr5− cells, which upon arrival seed metastases in which Lgr5+ cells appear. This plasticity can occur independently of stemness-inducing factors and is indispensable for long-term metastatic growth.
Various species of the intestinal microbiota have been associated with the development of colorectal cancer
, but it has not been demonstrated that bacteria have a direct role in the occurrence of ...oncogenic mutations. Escherichia coli can carry the pathogenicity island pks, which encodes a set of enzymes that synthesize colibactin
. This compound is believed to alkylate DNA on adenine residues
and induces double-strand breaks in cultured cells
. Here we expose human intestinal organoids to genotoxic pks
E. coli by repeated luminal injection over five months. Whole-genome sequencing of clonal organoids before and after this exposure revealed a distinct mutational signature that was absent from organoids injected with isogenic pks-mutant bacteria. The same mutational signature was detected in a subset of 5,876 human cancer genomes from two independent cohorts, predominantly in colorectal cancer. Our study describes a distinct mutational signature in colorectal cancer and implies that the underlying mutational process results directly from past exposure to bacteria carrying the colibactin-producing pks pathogenicity island.
The significance of cardiac stem cell (CSC) populations for cardiac regeneration remains disputed. Here, we apply the most direct definition of stem cell function (the ability to replace lost tissue ...through cell division) to interrogate the existence of CSCs. By single-cell mRNA sequencing and genetic lineage tracing using two Ki67 knockin mouse models, we map all proliferating cells and their progeny in homoeostatic and regenerating murine hearts. Cycling cardiomyocytes were only robustly observed in the early postnatal growth phase, while cycling cells in homoeostatic and damaged adultmyocardium represented various noncardiomyocyte cell types. Proliferative postdamage fibroblasts expressing follistatin-like protein 1 (FSTL1) closely resemble neonatal cardiac fibroblasts and form the fibrotic scar. Genetic deletion of Fstl1 in cardiac fibroblasts results in postdamage cardiac rupture. We find no evidence for the existence of a quiescent CSC population, for transdifferentiation of other cell types toward cardiomyocytes, or for proliferation of significant numbers of cardiomyocytes in response to cardiac injury.
Developmental gene expression is often controlled by distal regulatory DNA elements called enhancers. Distant enhancer action is restricted to structural chromosomal domains that are flanked by ...CTCF-associated boundaries and formed through cohesin chromatin loop extrusion. To better understand how enhancers, genes and CTCF boundaries together form structural domains and control expression, we used a bottom-up approach, building series of active regulatory landscapes in inactive chromatin. We demonstrate here that gene transcription levels and activity over time reduce with increased enhancer distance. The enhancer recruits cohesin to stimulate domain formation and engage flanking CTCF sites in loop formation. It requires cohesin exclusively for the activation of distant genes, not of proximal genes, with nearby CTCF boundaries supporting efficient long-range enhancer action. Our work supports a dual activity model for enhancers: its classic role of stimulating transcription initiation and elongation from target gene promoters and a role of recruiting cohesin for the creation of chromosomal domains, the engagement of CTCF sites in chromatin looping and the activation of distal target genes.
Abstract
Cycling Lgr5
+
stem cells fuel the rapid turnover of the adult intestinal epithelium. The existence of quiescent Lgr5
+
cells has been reported, while an alternative quiescent stem cell ...population is believed to reside at crypt position +4. Here, we generated a novel
Ki67
RFP
knock‐in allele that identifies dividing cells. Using
Lgr5‐
GFP
;Ki67
RFP
mice, we isolated crypt stem and progenitor cells with distinct Wnt signaling levels and cell cycle features and generated their molecular signature using microarrays. Stem cell potential of these populations was further characterized using the intestinal organoid culture. We found that Lgr5
high
stem cells are continuously in cell cycle, while a fraction of Lgr5
low
progenitors that reside predominantly at +4 position exit the cell cycle. Unlike fast dividing
CBC
s, Lgr5
low
Ki67
−
cells have lost their ability to initiate organoid cultures, are enriched in secretory differentiation factors, and resemble the Dll1 secretory precursors and the label‐retaining cells of Winton and colleagues. Our findings support the cycling stem cell hypothesis and highlight the cell cycle heterogeneity of early progenitors during lineage commitment.
Synopsis
image
A
Ki67
‐
RFP
knock‐in allele highlights the heterogeneity of Lgr5
+
stem‐ and progenitor cells based on
in vivo
cell cycle profiles.
Ki67
‐
RFP
allele allows identification and visualization of proliferating cells
in vivo
.
Lgr5
high
intestinal stem cells are continuously in the cell cycle.
Quiescent Lgr5
low
“+4” cells are secretory precursors.
Loci on mouse chromosome 2 have previously been associated with ethanol consumption. Here, we used a limited access choice paradigm in which mice consume large quantities of ethanol (2-3 g/kg/2 h) ...with a high preference (>80%). In addition, mouse chromosome substitution strains were used to further evaluate the contribution of chromosome 2 to ethanol consumption.
First, we compared the two parental inbred mouse strains, C57BL/6J and A/J, in the limited access choice paradigm for ethanol intake and ethanol preference, as well as for ethanol metabolism and taste sensitivity. Then, the effect of chromosome 2 substitution on these measures was determined. Compared with C57BL/6J mice, A/J and C57BL/6J-Chr 2/NaJ (CSS-2) mice showed profoundly reduced ethanol intake and preference. The strains were not different with regard to ethanol metabolism or taste sensitivity. Limited access ethanol consumption in F2 progeny derived from reciprocal C57BL/6J xCSS-2 and CSS-2 xC57BL/6J intercrosses and subsequent quantitative trait loci mapping identified two loci: one locus on chromosome 2 for ethanol intake and a separate locus on distal chromosome 2 for ethanol preference. This latter locus was dependent on the grandparental origin.
Using a limited access choice paradigm, we found that mouse chromosome 2 carries an allelic variant of a locus for ethanol intake and a distinct locus selective for ethanol preference. The heritability of alcoholism has been suggested to be parent-specific, perhaps resulting from genetic imprinting. Our findings suggest that grandparent-influenced vulnerability for ethanol consumption is conferred by genes on chromosome 2, providing important new leads to enhance our understanding of the heritability of alcoholism.
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