Bmi1 is expressed in vivo in intestinal stem cells

Abstract

Bmi1 plays an essential part in the self-renewal of hematopoietic and neural stem cells. To investigate its role in other adult stem cell populations, we generated a mouse expressing a tamoxifen-inducible Cre from the Bmi1 locus. We found that Bmi1 is expressed in discrete cells located near the bottom of crypts in the small intestine, predominantly four cells above the base of the crypt (+4 position). Over time, these cells proliferate, expand, self-renew and give rise to all the differentiated cell lineages of the small intestine epithelium. The induction of a stable form of b-catenin in these cells was sufficient to rapidly generate adenomas. Moreover, ablation of Bmi1 + cells using a Rosa26 conditional allele, expressing diphtheria toxin, led to crypt loss. These experiments identify Bmi1 as an intestinal stem cell marker in vivo. Unexpectedly, the distribution of Bmi1-expressing stem cells along the length of the small intestine suggested that mammals use more than one molecularly distinguishable adult stem cell subpopulation to maintain organ homeostasis.Adult somatic stem cells are defined by two major properties: the ability to generate more stem cells (self-renewal) and the ability to generate differentiated cell lineages 1 . To establish the presence of these two properties, the gold standard is to assess both of them in vivo and in vitro 2 . Several studies have shown in vitro that many tissues carry cells capable of selfrenewal and of giving rise to differentiated cell types. However, few experiments have clearly established in vivo self-renewal and multipotency of these cells over time 3,4 .To address this, we established a genetic fate-mapping system for stem cell populations in vivo. We chose as our target locus Bmi1, a gene already known to be involved in the selfrenewal of neuronal, hematopoietic and leukemic cells [5][6][7] . Bmi1 was first identified in a mouse proviral insertion screen for lymphomagenesis 8 . It is part of the Polycomb group gene family, and specifically a member of polycomb-repressing complex 1 (PRC1). PRC1 has an essential role in maintaining chromatin silencing 9, 10. Bmi1 −/− mice die before or near weaning from a defect in self-renewal of hematopoietic stem cells 11,12 . Our hypothesis was that PRC1 might be part of a general mechanism for maintaining self-renewal in various adult stem cell populations.We inserted a construct encoding a internal ribosome entry site (IRES)-Cre-estrogen receptor binding domain fusion (IRES-Cre-ER) into the 3′ untranslated region of Bmi1 ( Supplementary Fig. 1 To evaluate the repopulation kinetics of the crypt cells derived from Bmi1 + parents, we analyzed the small intestines of Bmi1 Cre-ER/+ ;Rosa26 LacZ/+ mice from day 2 to day 30 after tamoxifen induction, analyzing and following crypts on serial adjacent sections. Crypts with LacZ + cells presented a large variation in the number of stained cells. From day 2 to day 30, there was a continual increase in the number of cells labeled per crypt . Over time, the number of crypts th...

Identification of Lineage-Uncommitted, Long-Lived, Label-Retaining Cells in Healthy Human Esophagus and Stomach, and in Metaplastic Esophagus

“…In this regard recent genetic lineage-tracing studies in the mouse have provided new evidence to support the existence of both rapid and slowly cycling intestinal stem cells. 1 Furthermore murine LRCs are most commonly found at position +4 crypts of the small intestine, 27 where the putative stem cell marker Bmi-1 is expressed 28,29 however Bmi-1 was not expressed in Barrett's glands from our patients (unpublished observations) and there are insufficient numbers of LRCs to precisely determine their cell position. It is possible that two stem cell populations co-exist in the gut in an overlapping way, one rapidly cycling and the other slowly cycling and that these may be interconvertable 29,30 .…”

Attributes of adult stem cells

“…In the mouse small intestine, Bmi1-expressing cells can be localized by in situ hybridization as being located at cell position 4-5, counting from the bottom of the crypt, and lineage tracing from Bmi1-expressing cells suggests that these cells can also generate long-lived clones containing all the intestinal cell lineages [63].…”

“…CBCs are actively cycling cells -nevertheless, they appear to function as stem cells; using a tamoxifen-activatable Cre recombinase knocked into the Lgr5 locus, it has been possible to find long-lived clones derived from Lgr5-positive cells that contained all small intestinal lineages, likewise in the large intestine [4]. A similar strategy based on Bmi1-positive cells, also generated multilineage clones, but the Bmi1-positive cells were located at cell position 4-5 [63]. Thus, in the mouse small intestine at least, we appear to have two distinct stem cell populations, rapidly cycling Lgr5 + CBC stem cells and a more slowly cycling Bmi1 + stem cell population at positions 4-5.…”

Microenvironmental control of stem cell fate in intestinal homeostasis and disease

“…Several cell markers with expression patterns that overlie this position have been described 12, 13, 14. Using a tamoxifen‐inducible Bmi1 Cre recombinase mouse, Sangiorgi et al demonstrated lineage tracing from cells at the +4 position 12.…”