Reviews in Basic and Clinical GastroenterologyLeucine-Rich Repeat-Containing G-Protein-Coupled Receptors as Markers of Adult Stem Cells
Section snippets
Lgr5 Is a Wnt Target Gene in Colon Cancer
The Wnt signaling pathway regulates the proliferative activity of intestinal crypt cells.4, 5, 6 It is therefore not surprising that mutations that activate the Wnt pathway, most notably through loss of its negative regulator, adenomatous polyposis coli (APC), have been associated with many forms of colon cancer.7, 8 Wnt signaling ultimately induces transcription of genes via transcription factors of the T-cell factor (Tcf) family. The genetic programs that are activated by Wnt/Tcf signaling
The Lgr4, Lgr5, and Lgr6 Receptors
In 1998, Hsu et al16 cloned 2 molecules that were related to the hormone receptors for thyroid-stimulating hormone (TSH), follicle-stimulating hormone (FSH), and leutinizing hormone (LH). These receptors belong to the large, G-protein–coupled, 7-transmembrane (7TM) family of proteins. They are unique in that they have a large N-terminal extracellular (ecto-) domain that contains a series of leucine-rich repeats. In the LH-, FSH-, and TSH-receptor molecules, the ectodomain is crucial for binding
Ligands for Lgr4, Lgr5, and Lgr6
To fully understand the signaling functions of the Lgr4, Lgr5, and Lgr6 orphan G-protein-coupled receptors (GPCRs), it is important to identify their elusive ligands. Some predictions can be made based on the ligands of related receptors. Phylogenetic analysis indicates that there are 3 Lgr subgroups: the LH/FSH/TSH glycoprotein hormone receptors; Lgr4, Lgr5, and Lgr6; and Lgr7 and Lgr8 (Figure 2B). The ligands of the first subgroup (LH, FSH, and TSH) are defined structurally by the presence of
Genetic Analysis of Lgr4 Function in Mice
Lgr4 has been better studied than Lgr5 or 6. Several mutant alleles of mouse Lgr4 have been generated and analyzed and the Lgr4 expression pattern has been well characterized35, 36 (Figure 3). A gene-trapping screen identified a mouse line (Lgr4Gt(pGTOTMpfs)1Wcs) in which the Lgr4 gene was disrupted by a gene trap vector that expressed a bacterial β-galactosidase LacZ fusion protein and placental alkaline phosphatase (Figure 3A).37 Van Schoore et al36 investigated LacZ and placental alkaline
Lgr5 Function
Little was known about mammalian Lgr5 before 2007. Lgr5/Gpr49 is a Wnt target gene as well as a cancer gene; it was on the original list of Wnt/Tcf4 targets that are active in colorectal cancers9 and is overexpressed in tumors of the ovary and liver, likely because of the mutational activation of the Wnt pathway in these tumors.49, 50, 51Lgr5 expression was observed in basal cell carcinomas52 and in healthy cyclic endometrium.53
The phenotype of a mouse knockout for Lgr5 (Lgr5tm1Ah) was
Lgr5 Marks Stem Cells in the Intestine
The crypts of the small intestine contain functional stem cells. Although researchers agree that every crypt contains 4–6 independent stem cells, there is disagreement about their exact identity.56 The stem cells are believed to reside at position +4 (on average), relative to the crypt bottom, with positions 1–3 occupied by the terminally differentiated Paneth cells. Potten et al57, 58 provided experimental support for the +4 stem cell model, reporting that radiation-sensitive, label-retaining
Lgr5 Marks Stem Cells in the Stomach
The stomach is similar to the intestine; it is of common endodermal origin and has a constantly renewing epithelium. The equivalent of the intestinal crypt is called the gastric gland (or glandular unit) in the stomach epithelium. Several of these glands feed into a pit that opens out onto the surface epithelium. The structure and composition of these gastric units varies in different anatomic regions of the stomach.66 In the pyloric region (antrum), each gastric unit is composed of several
Lgr5 Marks Stem Cells in the Hair Follicle
The control of self-renewal in cells of the intestinal crypts is similar to that of hair follicles; both processes are regulated by Wnt signaling.81 Although the interfollicular epidermis undergoes constant self-renewal, the hair follicles go through cyclic phases of growth, involution, and rest.82 In mice, hair follicle stem cells reside in the follicle bulge and are characterized by expression of the cell-surface marker CD34 and cytokeratin 15.83, 84, 85, 86 These stem cells are believed to
Lgr6 as a Marker of Adult Stem Cells in Skin
Because Lgr5 appeared to be expressed by a variety of unrelated adult stem cells, we obtained mice with LacZ- or GFP-ires-CreERT2 inserted at Lgr4 and Lgr6, to identify cells that express these genes and to assess their stem cell potential via lineage tracing. Lgr4 does not seem to be a marker of stem cells—it is expressed more broadly and in many proliferative compartments36 (and Haegebarth et al, unpublished data). In contrast, Lgr6 expression in adult Lgr6-LacZLacZ knock-in mice (Figure 3F)
Refining Experimentation on Adult Stem Cells
Lgr5 and 6 are unique markers in that they can be used to discriminate stem cells from their immediate, transit-amplifying daughters. Other candidate markers of intestinal stem cells, such as Musashi94 or CD133,95, 96, 97 typically are expressed in a shallower gradient along the crypt axis and a consequently broader pattern. Similarly, doublecortin and calcium/calmodulin-dependent kinase-like 1 (DCAMKL-1) is expressed on a rare population of terminally differentiated cells, called Tuft cells,
Future Directions
In all tissues in which Lgr5- and Lgr6-expressing cells have been studied by lineage tracing, these cells have been found to represent stem cells. Most attention has been restricted to tissues that display a spontaneous, high level of self-renewal activity. In many organs, expression of the 2 markers is observed during development and/or upon injury, but does not occur in the adult under normal conditions. These tissues typically have very low rates of spontaneous self-renewal (eg, pancreas,
Acknowledgments
The authors thank Hugo Snippert for Lgr6 tracing figures; and Gilbert Vassart, Aaron Hsueh, and laboratory members for critical review of the manuscript.
Both authors contributed equally to the design and writing of this review.
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Conflicts of interest The authors disclose no conflicts.