SnapShot: The Intestinal Crypt

1198

Cell 152, February 28, 2013 ©2013 Elsevier Inc.

DOI http://dx.doi.org/10.1016/j.cell.2013.02.030

See online version for legend and references.

PEYER’S PATCH

Lymphoid cells

Paneth cell precursors Quiescent cells Regeneration of ISCs

Label retaining cells (LRCs)

Tissue renewal

Intestinal stem cells (ISCs)

Antigen sampling

M cells

Hormone production

Enteroendocrine cells

Mucus secretion

Goblet cells

Opioids release Prostanoids production Other?

Tuft cells

Innate immune response Stem cell niche maintenance

Paneth cells

Proliferation Lineage commitment

Ileum

CRYPT

Cell amplification

Migration

Anoikis

The small intestine

Jejunum

Nutrients

Prox. colon

Caecum

Represses Ileal gene program

Cdx2

Proximodistal organization

Gata4

Duodenum

Transient amplifying cells

Nutrients absorption

Enterocytes

Stem cell niche

Transient-amplifying compartment

Differentiated cell compartment

Stomach

Represses stomach gene program

VILLUS

Dist. colon

?

Spi-B

(Lgr5+)

ISCs

Da

ma

Damage

Absorptive progenitor

Notch

Hes1

Elf3 Cdx2+Hnf4

Enterocyte

STROMAL CELL

BMP

GREM

BMPRII

Proliferation, block differentiation: MYC Regulation of WNT signaling: ZNRF3/RNF43, LGR5 Cell positioning: EPHB2, EPHB3 Self-renewal: ASCL2

BMPRI

Pax4/6 Nkx2.2 Ngn3

NeuroD

Endocrine

?

WNT

Neutral competition in the ISC niche

WNT

ZNRF3/ RNF43

ISC (CBC)

NOTCH-IC

TIME

WNT

?

EGF

LRIG1

?

ERBB

DLL1/4

NOTCH

WNT

TGFα

PA N E T H CELL

Crypts inevitably drift toward clonality

Growth of surviving clones and extinction of neighboring clones

Symmetric stem cell divisions

Restricted niche for stem cells

Paneth cells Lgr5+ stem cells

Intestinal stem cells at the crypt base

Proliferation

LRP6 FRIZZLED

β-cat

β-cat

RSPO

LGR4/5

TCF

CSL

Signaling in ISCs

Paneth cell

Gfi1 Sox9 Spdef Math1 +WNT

(Lgr5+)

LRC

ge

Math1

Elf3 Spdef Gfi1

Goblet

Secretory progenitor

Dll1/4

?

Tuft

Inhibition of the secretory program: HES1 Cell positioning: EPHB2 Silencing of cell-cycle inhibitors (p27kip, p57kip) Repression of expression of Dll1/4

RankL

M cell

Cell hierarchy and lineage specification

Hans Clevers1 and Eduard Batlle2,3 1 Hubrecht Institute, KNAW and University Medical Centre Utrecht, Uppsalalaan 8, 3584CT Utrecht, the Netherlands 2 Institute for Research in Biomedicine (IRB Barcelona), 08028 Barcelona, Spain 3 Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona, Spain

SnapShot: The Intestinal Crypt

SnapShot: The Intestinal Crypt Hans Clevers1 and Eduard Batlle2,3 1 Hubrecht Institute, KNAW and University Medical Centre Utrecht, Uppsalalaan 8, 3584CT Utrecht, the Netherlands 2 Institute for Research in Biomedicine (IRB Barcelona), 08028 Barcelona, Spain 3 Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona, Spain Organization of the Small Intestine and the Colon The inner surface of the intestinal tube is lined by a simple epithelium, which displays distinct morphologies along the proximo-distal axis. The proximal region, the small intestine (duodenum, jejunum, and ileum), is arranged in invaginations (crypts) intercalated with finger-like protrusions (villus) that represent units specialized in the absorption of micronutrients. The colon consists only of crypts and is largely dedicated to the compaction of stool. The identity of each of these segments is, in part, specified through the expression of the homeotic transcription factor Cdx2, which represses the expression of genes characteristic of the stomach, and the zinc finger Gata4, which confers proximal identity to the duodenum and ileum. Despite their distinctive morphologies, epithelial cell types of both the small intestine and colon are organized following a bottom-to-top axis into three compartments: the stem cell compartment that is located at the crypt base, the transient-amplifying (TA) compartment that occupies the middle portion of the crypts, and the differentiation zone, which expands from the top third of the crypt and the surface epithelium to the tip of the villus. Renewal of the Epithelial Layer and Lineage Specification Six differentiated cell types populate the intestinal epithelium. The most abundant cells are the enterocytes (i.e., adsorptive cells) and goblet cells (i.e., mucosecreting cells). Other rare secretory cell types are scattered throughout the epithelium; they are enteroendocrine cells, which produce a diverse array of hormones, and the tuft cells, which are believed to secrete prostanoids. Microfold cells (M cells) are specialized epithelial cells situated over Peyer’s patches (PP) that transport antigens into intraepithelial pockets accessed by antigen-presenting cells. Finally, Paneth cells reside at the crypt base and perform a dual role; they secrete antimicrobial substances but also nurture the intestinal stem cell (ISC) population. All differentiated cell types in the intestinal epithelium are short lived. Enterocytes, goblet cells, and the other secretory lineages are born in the crypts and follow an upward migratory flow that carries them to the tip of the villi in ~1 week. At this location, they are extruded into the lumen. Paneth cells are the exception and are retained at the base of the crypts, where they live for 6–8 weeks. Renewal of the epithelial layer is sustained throughout life by a small number of ISCs (n = 10–15 per crypt), which are located at the bottommost positions intermingled with Paneth cells. ISCs proliferate with a rate of approximately one division per day. Their progeny is amplified through a series of very rapid divisions (about one division every 12 hr). While these TA cells migrate upward, they become progressively committed toward one of six lineages that are present in the intestine. The activity of the bHLH transcription factor Math1 commits precursor cells to a secretory phenotype. Math1 also promotes elevated levels of delta-like ligands (Dll1/Dll4) in secretory precursors. In contrast, expression of Math1 is repressed in enterocytes by the Notch downstream effector Hes1. As a result of lateral inhibition, differential Notch activity in adjacent precursors operates as a binary switch to specify absorptive versus secretory cell types. Following commitment, a complex cascade of transcription factors drives the differentiation of the secretory precursor into distinct mature cell types (goblet, enteroendocrine, tuft, or Paneth cells). Differentiation of M cells requires the transcription factor Spi-B, the expression of which is switched on downstream of RANK signaling. RANK-L, the ligand for RANK receptor, is secreted by stromal cells present at Peyer’s patches. Quiescent versus Proliferative ISCs In homeostasis, Lgr5+ ISCs generate all cell types present in the epithelium (Barker et al., 2007; Sato et al., 2009). An independent class of quiescent ISCs marked by Bmi1, Lrig1, Tert, and Hopx has been proposed to serve as “reserve” stem cells. A recent study provides a simple view of the connection between these two stem cell types (Buczacki et al., 2013). The study shows that a small subset of Lgr5+ cells can enter a quiescent state. These label-retaining cells (LRCs) localize around crypt position +3. Lgr5+ LRCs express most of the markers previously postulated for the quiescent ISC population, but also several Paneth-cell-specific genes. These LRCs are not ISCs but, rather, transient precursors of Paneth cells, as they are short lived under homeostatic conditions (2–3 weeks) and they all differentiate toward mature Paneth cells. However, upon loss of the ISC pool, they can revert to become cycling Lrg5+ ISCs. Dedifferentiation of Dll1+-committed secretory precursors upon irradiation followed by regeneration of the ISC pool has also been demonstrated (van Es et al., 2012). Neutral Competition in the ISC Niche An intestinal crypt contains about 14 equal ISCs that all divide each day. Their dynamics are consistent with a model in which the resident ISCs double their numbers each day and stochastically adopt either stem or TA fates. Thus, ISCs divide symmetrically while competing for a niche of limited size. As a consequence, their turnover follows a pattern of neutral drift dynamics and crypts tend toward clonality within a period of 1–6 months. Therefore, ISCs persist for life as a population, yet only the lineage of one particular ISC is present in each crypt at any given time (Snippert et al., 2010). Signaling in ISCs ISCs are specified by high levels of WNT signaling in the crypts. R-spondin binds to LGR4/LGR5 receptors and potentiates WNT signals in ISCs (de Lau et al., 2011). On the contrary, the activity of Znfr3/Rnf43 negatively controls WNT signals in the ISC pool by ubiquitinating Frizzled receptors (Koo et al., 2012). Paneth cells secrete the WNT3 ligand constitutively, but an additional Wnt source also exists in the surrounding stroma. Notch signaling mainly acts by inhibiting the secretory fate in ISCs (Pellegrinet et al., 2011). Notch ligands (Dll1 and Dll4) are expressed by surrounding secretory cells, including the Paneth cells. Mitogenic stimuli regulate the size of the proliferative compartment. Lrig1, a marker for stem and early TA cells, acts as a negative regulator of receptor tyrosine kinase (RTK) activity (Wong et al., 2012). BMP signaling inhibits the stem cell fate in the intestine (Haramis et al., 2004) probably by antagonizing WNT signaling. BMPs are mainly expressed by stromal cells that surround the epithelium, whereas ISCs are protected from their action by the presence of local inhibitors, including Gremlin. References Barker, N., van Es, J.H., Kuipers, J., Kujala, P., van den Born, M., Cozijnsen, M., Haegebarth, A., Korving, J., Begthel, H., Peters, P.J., et al. (2007). Identification of stem cells in small intestine and colon by marker gene Lgr5. Nature 449, 1003–1007. Buczacki, S.J.A., Zecchini, H.I., Nicholson, A.M., Russell, R., Vermeulen, L., Kemp, R., and Winton, D.J. (2013). The intestinal label-retaining cell expresses Lgr5 and is a committed secretory precursor. Nature. Published online February 27, 2013. http://dx.doi.org/10.1038/nature11965. de Lau, W., Barker, N., Low, T.Y., Koo, B.K., Li, V.S., Teunissen, H., Kujala, P., Haegebarth, A., Peters, P.J., van de Wetering, M., et al. (2011). Lgr5 homologues associate with Wnt receptors and mediate R-spondin signalling. Nature 476, 293–297. Haramis, A.P., Begthel, H., van den Born, M., van Es, J., Jonkheer, S., Offerhaus, G.J., and Clevers, H. (2004). De novo crypt formation and juvenile polyposis on BMP inhibition in mouse intestine. Science 303, 1684–1686. Koo, B.K., Spit, M., Jordens, I., Low, T.Y., Stange, D.E., van de Wetering, M., van Es, J.H., Mohammed, S., Heck, A.J., Maurice, M.M., and Clevers, H. (2012). Tumour suppressor RNF43 is a stem-cell E3 ligase that induces endocytosis of Wnt receptors. Nature 488, 665–669.

1198.e1  Cell 152, February 28, 2013 ©2013 Elsevier Inc.  DOI http://dx.doi.org/10.1016/j.cell.2013.02.030

SnapShot: The Intestinal Crypt Hans Clevers1 and Eduard Batlle2,3 1 Hubrecht Institute, KNAW and University Medical Centre Utrecht, Uppsalalaan 8, 3584CT Utrecht, the Netherlands 2 Institute for Research in Biomedicine (IRB Barcelona), 08028 Barcelona, Spain 3 Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona, Spain

Pellegrinet, L., Rodilla, V., Liu, Z., Chen, S., Koch, U., Espinosa, L., Kaestner, K.H., Kopan, R., Lewis, J., and Radtke, F. (2011). Dll1- and dll4-mediated notch signaling are required for homeostasis of intestinal stem cells. Gastroenterology 140, 1230–1240.e1–e7. Sato, T., Vries, R.G., Snippert, H.J., van de Wetering, M., Barker, N., Stange, D.E., van Es, J.H., Abo, A., Kujala, P., Peters, P.J., and Clevers, H. (2009). Single Lgr5 stem cells build crypt-villus structures in vitro without a mesenchymal niche. Nature 459, 262–265. Snippert, H.J., van der Flier, L.G., Sato, T., van Es, J.H., van den Born, M., Kroon-Veenboer, C., Barker, N., Klein, A.M., van Rheenen, J., Simons, B.D., and Clevers, H. (2010). Intestinal crypt homeostasis results from neutral competition between symmetrically dividing Lgr5 stem cells. Cell 143, 134–144. van Es, J.H., Sato, T., van de Wetering, M., Lyubimova, A., Nee, A.N., Gregorieff, A., Sasaki, N., Zeinstra, L., van den Born, M., Korving, J., et al. (2012). Dll1+ secretory progenitor cells revert to stem cells upon crypt damage. Nat. Cell Biol. 14, 1099–1104. Wong, V.W., Stange, D.E., Page, M.E., Buczacki, S., Wabik, A., Itami, S., van de Wetering, M., Poulsom, R., Wright, N.A., Trotter, M.W., et al. (2012). Lrig1 controls intestinal stem-cell homeostasis by negative regulation of ErbB signalling. Nat. Cell Biol. 14, 401–408.

1198.e2  Cell 152, February 28, 2013 ©2013 Elsevier Inc.  DOI http://dx.doi.org/10.1016/j.cell.2013.02.030