Nod-like Receptors Have a Grip on Stem Cells
Authors: Jörg H. Fritz
Journal: Cell Host & Microbe (2014)

Two reports in this issue of Cell Host & Microbe establish that Nod-like receptor proteins NOD1 and NOD2 regulate stem cell function. Burberry et al., 2014Burberry A. Zeng M.Y. Ding L. Wicks I. Inohara N. Morrison S.J. Núñez G. Cell Host Microbe. 2014; 15 (this issue): 779-791Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar demonstrate that NOD1 and NOD2 synergize with TLRs to mobilize hematopoietic stem cells. Nigro et al., 2014Nigro G. Rossi R. Commere P.-H. Jay P. Sansonetti P.J. Cell Host Microbe. 2014; 15 (this issue): 792-798Abstract Full Text Full Text PDF PubMed Scopus (178) Google Scholar report that NOD2 provides cytoprotection to intestinal stem cells. Two reports in this issue of Cell Host & Microbe establish that Nod-like receptor proteins NOD1 and NOD2 regulate stem cell function. Burberry et al., 2014Burberry A. Zeng M.Y. Ding L. Wicks I. Inohara N. Morrison S.J. Núñez G. Cell Host Microbe. 2014; 15 (this issue): 779-791Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar demonstrate that NOD1 and NOD2 synergize with TLRs to mobilize hematopoietic stem cells. Nigro et al., 2014Nigro G. Rossi R. Commere P.-H. Jay P. Sansonetti P.J. Cell Host Microbe. 2014; 15 (this issue): 792-798Abstract Full Text Full Text PDF PubMed Scopus (178) Google Scholar report that NOD2 provides cytoprotection to intestinal stem cells. The host-encoded pattern-recognition molecules (PRMs) nucleotide-binding oligomerization domain 1 (NOD1, also referred to as NRLC1) and NOD2 (NLRC2), which belong to the Nod-like receptor (NLR) protein family, are known to sense distinct bacterial peptidoglycan (PGN) fragments derived from pathogens, commensals, and pathobionts to instruct innate and adaptive antimicrobial immunity and regulate mucosal immune homeostasis (Fritz et al., 2006Fritz J.H. Ferrero R.L. Philpott D.J. Girardin S.E. Nat. Immunol. 2006; 7: 1250-1257Crossref PubMed Scopus (707) Google Scholar). Now, two recent reports from Nigro et al., 2014Nigro G. Rossi R. Commere P.-H. Jay P. Sansonetti P.J. Cell Host Microbe. 2014; 15 (this issue): 792-798Abstract Full Text Full Text PDF PubMed Scopus (178) Google Scholar and Burberry et al., 2014Burberry A. Zeng M.Y. Ding L. Wicks I. Inohara N. Morrison S.J. Núñez G. Cell Host Microbe. 2014; 15 (this issue): 779-791Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar published in this issue of Cell Host & Microbe extend our knowledge of how these NLR proteins instruct immunity by demonstrating that PGN sensing by NOD1 and NOD2 regulate key biological functions of hematopoietic stem cells (HSCs) and intestinal stem cells (Figure 1). During infection and inflammation, immune cells are in high demand and are quickly consumed at the sites of need. While adaptive immune cells have high proliferative capacity, innate immune cells are mostly postmitotic and need to be rapidly replenished from bone marrow (BM) hematopoietic stem and progenitor cells. It is estimated that in an adult human, ∼3 × 105 erythrocytes and 3 × 104 white blood cells are produced per second at steady state, and upon infectious challenge the cellular production can increase several-fold over baseline levels. Obviously, such high cellular throughput requires a tight hierarchical control that is organized with the HSCs at the top. The majority of HSCs reside in the BM within the stem cell niche, while a smaller subset of HSCs reside in the spleen and serve as a site for hematopoiesis during embryogenesis and periods of duress. Although niche support is essential for their maintenance, HSCs traffic through the bloodstream and through peripheral tissues and have the potential to serve as a local source for cell production. While much is known about HSC regulation and activity under homeostatic conditions, how HSC throughput is regulated during infection and inflammation and the contributions of distinct PRMs is less clear. Recent work utilizing Toll-like receptor (TLR)-deficient settings and application of purified TLR agonists has provided evidence that indirect as well as direct activation of HSCs by TLR agonists is critical for demand-adapted hematopoiesis during microbial encounter (Takizawa et al., 2012Takizawa H. Boettcher S. Manz M.G. Blood. 2012; 119: 2991-3002Crossref PubMed Scopus (275) Google Scholar). In settings of systemic infection, microbe-associated molecular patterns (MAMPs) activating TLR signaling are elevated in circulation and reach the BM, where they directly act on TLR-expressing HSCs and instruct enhanced generation of a lineage on demand. Upon encounter of invading pathogens in peripheral tissues, microbe-mediated triggering of TLR on HSCs induces migration arrest and promotes HSC proliferation and differentiation to ensure local supply of myeloid effector cells. During the course of an immune response, when certain lineages are in high demand at specific sites, TLR ligation on lineage-committed progenitors in the BM is thought to cause mobilization from the BM and recruitment to these sites, where they proliferate and differentiate in response to local pathogens to replenish immune effector cells (Yáñez et al., 2013Yáñez A. Goodridge H.S. Gozalbo D. Gil M.L. Eur. J. Immunol. 2013; 43: 2526-2533Crossref PubMed Scopus (55) Google Scholar). As Escherichia coli is commonly isolated from the blood of humans suffering bacteremia, Burberry et al., 2014Burberry A. Zeng M.Y. Ding L. Wicks I. Inohara N. Morrison S.J. Núñez G. Cell Host Microbe. 2014; 15 (this issue): 779-791Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar infected rodents with this Gram-negative bacterium to study the mechanisms by which systemic bacterial infection influences localization and function of HSCs. The authors demonstrate that infection with E. coli increases the number of HSCs in the spleen and moderately reduces HSCs in the BM. Importantly, splenic HSC accumulation was greatly diminished in TLR4−/− mice and partially diminished in animals lacking the adaptor protein receptor-interacting serine-threonine kinase 2 (RIPK2), an essential downstream component of both NOD1 and NOD2 signaling, suggesting that during infection the synergistic activation of TLR4 and NOD1/NOD2 is required. Interestingly, administration of the TLR4 agonist lipopolysaccharide (LPS) did not induce splenic HSC accumulation, while coadministration of LPS and NOD1 agonists was sufficient, highlighting a previously unappreciated role for NLR signaling in promoting extramedullary hematopoiesis. Activation of TLR4 and NOD1 synergistically induced granulocyte colony-stimulating factor (G-CSF), which was required for splenic HSC accumulation. Mechanistically, Burberry et al. further show in a set of elegant experiments that TLR4 and RIPK2 expression in radio-resistant cells was required for splenic HSC accumulation, while TLR4 expression on HSCs was dispensable. These observations are in agreement with a recent report demonstrating that LPS-induced G-CSF-mediated emergency myelopoiesis depends on TLR4-expressing nonhematopoietic cells (Boettcher et al., 2012Boettcher S. Ziegler P. Schmid M.A. Takizawa H. van Rooijen N. Kopf M. Heikenwalder M. Manz M.G. J. Immunol. 2012; 188: 5824-5828Crossref PubMed Scopus (106) Google Scholar). Future experiments will be required to define the nonhematopoietic cell type(s) that produce G-CSF upon PRM stimulation. Whether the accumulation of splenic HSCs that occurs following infection is due to expansion of resident HSCs or mobilization of HSCs from the BM also remains to be determined. In support of mobilization, Burberry et al., 2014Burberry A. Zeng M.Y. Ding L. Wicks I. Inohara N. Morrison S.J. Núñez G. Cell Host Microbe. 2014; 15 (this issue): 779-791Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar observed that less than 5% of splenic HSCs were actively cycling at a time when their numbers were rapidly increasing. As such it remains to be established whether the increase of splenic HSCs is due to an active recruitment process or due to inhibition of steady-state egress. Another possibility is that infection and/or PRM-mediated inflammation regulates the BM stem cell niche. CXCL12 is a chemokine critical for retention of HSC in the BM niche, and downregulation of CXCL12 is a common feature of G-CSF-mediated mobilization. Indeed, CXCL12 expression in BM was reduced after E. coli infection or dual stimulation of TLR4 and NOD1, suggesting that the PRM activation of stromal cells might regulate G-CSF and CXCL12 levels in parallel. As distinct cellular sources and niches in BM regulate HSC residency and function (Frenette et al., 2013Frenette P.S. Pinho S. Lucas D. Scheiermann C. Annu. Rev. Immunol. 2013; 31: 285-316Crossref PubMed Scopus (335) Google Scholar), which specific cell type(s) are regulated by distinct PRMs remains to be examined. Functionally, Burberry et al. further demonstrated that infection-induced accumulation of splenic HSCs and progenitor cells gives rise to neutrophils and monocytes, which contribute to limit secondary infection. Therefore, it is likely that the accumulation of HSCs in the spleen provides an additional source for the generation of effector cells during infection and inflammation, highlighting a previously unappreciated role for the synergistic function of TLR and NLR signaling. Recognition of bacterial PGN by NOD1 and NOD2 is also critical for the regulation of intestinal immune homeostasis. The intestinal microbiota protects the host against pathogenic microorganisms, increases its metabolic capacities, and shapes maturation of the local and systemic innate and adaptive immune system. The beneficial effects of microbiota on gut homeostasis occur in part through direct sensing of bacterial products by PRM. NOD1- and NOD2-mediated PGN recognition of intestinal bacteria has been shown to be critical for intestinal lymphoid tissue formation (Bouskra et al., 2008Bouskra D. Brézillon C. Bérard M. Werts C. Varona R. Boneca I.G. Eberl G. Nature. 2008; 456: 507-510Crossref PubMed Scopus (791) Google Scholar), and polymorphisms in NOD2 are linked to Crohn's disease (CD) (Rosenstiel et al., 2009Rosenstiel P. Sina C. Franke A. Schreiber S. Semin. Immunol. 2009; 21: 334-345Crossref PubMed Scopus (67) Google Scholar). Despite the strong association between CD and causative NOD2 variants, a comprehensive picture of how NOD2 contributes to disease pathogenesis has not yet emerged. NOD2 is broadly expressed in myeloid cells and at lower levels in lymphoid as well as nonhematopoietic cells, including intestinal epithelial cells (IECs). NOD2 is activated by N-acetyl muramyl dipeptide (MDP), a component of the PGN of Gram-negative and Gram-positive bacteria. Common CD-associated NOD2 variants abrogate RIPK2 binding and subsequent downstream signaling, suggesting a hypomorphic function of CD-associated NOD2 variants. However, some polymorphisms have been suggested to encode a gain-of-function aspect by actively suppressing transcription of the immune suppressive cytokine interleukin (IL)-10. Furthermore, Nod2-deficient mice exhibit decreased expression of specific antimicrobial α-defensins by epithelial cells, and patients with CD-associated NOD2 variants expressed decreased levels of α-defensins, suggesting that altered NOD2 function in CD leads to a reduction in the antimicrobial and anti-inflammatory response, contributing to chronic intestinal inflammation (Fritz et al., 2011Fritz T. Niederreiter L. Adolph T. Blumberg R.S. Kaser A. Gut. 2011; 60: 1580-1588Crossref PubMed Scopus (163) Google Scholar). A report published in this issue by Nigro et al., 2014Nigro G. Rossi R. Commere P.-H. Jay P. Sansonetti P.J. Cell Host Microbe. 2014; 15 (this issue): 792-798Abstract Full Text Full Text PDF PubMed Scopus (178) Google Scholar highlights a previously unrecognized role for NOD2 in intestinal homeostasis. Utilizing intestinal organoid culture systems (Barker et al., 2012Barker N. van Oudenaarden A. Clevers H. Cell Stem Cell. 2012; 11: 452-460Abstract Full Text Full Text PDF PubMed Scopus (237) Google Scholar), the authors tested the specific role of MAMPs activating TLR or NLR to monitor their effects on the yield and development of organoids. While stimulations with agonists activating TLR2, TLR4, TLR5, TLR9, and NOD1 had no influence on organoid growth, selective stimulation of NOD2 yielded an increase of organoids. As no change in the proliferative capacity of crypt cells was observed, Nigro et al. hypothesized that in the presence of MDP, more crypt and/or intestinal stem cells could survive and generate organoids. Indeed, cultures of Lgr5+ intestinal stem cells only or Lgr5+ intestinal stem cells cocultured with paneth cells yielded enhanced organoid growth upon stimulation with the NOD2 agonist. As NOD2 expression was found to be high in Lgr5+ intestinal stem cells but low in paneth cells, the authors reasoned that enhanced organoid growth results from direct stimulation of stem cells by MDP exerting cytoprotective effects. To substantiate their hypothesis, Nigro et al. reduced the microbiota content of mice by antibiotic treatment and administered doxorubicin to induce oxidative stress and injury in intestinal epithelial stem cells in the absence or presence of MDP. Importantly, the reduced frequency of proliferating crypt cells upon doxorubicin treatment could be reversed by MDP administration, and the effect was NOD2 dependent. Moreover, doxorubicin treatment led to increased cell death in NOD2−/− animals, demonstrating that NOD2-mediated PGN recognition by intestinal stem cells exerts a cytoprotective function. Future experiments will have to decipher if NOD2 expression solely in intestinal epithelial cells is responsible for the observed function or whether other NOD2-expressing cells, such as mesenchymal and/or hematopoietic cells, also play a role. As the NOD2-mediated cytoprotective effect may have major positive medical benefits, studies detailing the underpinning molecular mechanisms are of utmost importance. Infection Mobilizes Hematopoietic Stem Cells through Cooperative NOD-like Receptor and Toll-like Receptor SignalingBurberry et al.Cell Host & MicrobeMay 29, 2014In BriefMechanisms underlying hematopoietic stem cell (HSC) and progenitor mobilization during infection are undefined. Burberry et al. report that during systemic E. coli infection, NOD1 and TLR4 signaling mediate HSC mobilization to the spleen by inducing G-CSF in nonhematopoietic cells. The mobilized HSCs give rise to myeloid cells that combat infection. Full-Text PDF Open ArchiveThe Cytosolic Bacterial Peptidoglycan Sensor Nod2 Affords Stem Cell Protection and Links Microbes to Gut Epithelial RegenerationNigro et al.Cell Host & MicrobeMay 29, 2014In BriefThe intestinal crypt is the site of epithelial regeneration and of potential interactions between microbiota products and stem cells. Nigro et al. show that intestinal Lgr5+ stem cells express the cytosolic innate immune sensor Nod2, whose stimulation by the bacterial ligand muramyl-dipeptide affords potent cytoprotection to the stem cells. Full-Text PDF Open Archive