Tissue homeostasis and aging: new insight from the fly intestine

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ScienceDirect Tissue homeostasis and aging: new insight from the fly intestine Louis Gervais and Allison J Bardin Adult somatic stem cells facilitate tissue homeostasis throughout the life of the organism. The mechanisms controlling stem cell activity are under intense scrutiny, with the aims of elucidating how they mediate tissue homeostasis, contribute to age-related decline of adult tissues, and promote tumorigenesis. Recently, the use of model systems such as the Drosophila intestine has enriched our understanding of how stem cells integrate local and systemic signals to maintain tissue and organs function in physiological conditions of homeostasis or after damage. Here we highlight recent advances made in this model allowing a better understanding of stem cell lineage decisions, their regulation by epithelial and intra-organ cues, and their altered activity during aging.

Address Institut Curie, PSL Research University, CNRS UMR 3215, INSERM U934, Stem Cells and Tissue Homeostasis Group, Paris, France Corresponding author: Bardin, Allison J ([email protected])

Current Opinion in Cell Biology 2017, 48:97–105 This review comes from a themed issue on Cell Dynamics Edited by Eugenia Piddini and Helen McNeill For a complete overview see the Issue and the Editorial Available online 16th July 2017 http://dx.doi.org/10.1016/j.ceb.2017.06.005 0955-0674/ã 2017 Elsevier Ltd. All rights reserved

Introduction The activity of adult stem cells (SCs) needs to be closely regulated in order to compensate for cell loss according to the requirements of the tissue. Defects in the control of SC activity are associated with both age related decline of the tissue and cancer [1,2]. 10 years since the initial characterization of midgut intestinal stem cells (ISCs) in Drosophila [3,4], the fly gut has provided important insight into the processes controlling normal tissue homeostasis and associated tissue dysfunction. Here, we describe recent advances in our knowledge of Drosophila ISC cell fate lineage decisions, local and longdistance control of ISC proliferative responses, and aging associated changes in the fly intestine. The mammalian and Drosophila intestines share many similarities in terms of organization, biological function, www.sciencedirect.com

cellular composition, and maintenance by ISCs. The fly digestive tract is composed of the foregut, the midgut, and the hindgut, the differences of which are reviewed elsewhere [5]. Here, we will focus exclusively on the midgut, which can be divided into 10-14 subregions based on gene expression, structure, and function [6,7] though sharing similar ISC lineages and cell fate decisions.

Mechanisms of cell fate acquisition in the ISC lineage Recent updates on the ISC lineage

Under normal conditions of renewal, ISCs divide primarily asymmetrically, leading to the formation of an ISC and an enteroblast (EB), which can further differentiate into either an enterocyte (EC) or enteroendocrine cell (EE) [3,4]. While it is clear that ISCs can produce EC and EE cells, recent findings support the notion that EC and EE cells originate from a separate progenitors differing in the activation of the Notch pathway; whereas Notch is essential to promote EC fate, it must be kept off or at low levels for EE fate acquisition [8,9,10]. Our recent work has shown that the Notch inhibitor, Numb, acts at the top of a hierarchy to promote EE fate [11]. Nevertheless, how, exactly, an EE cell is specified is still somewhat obscure. Lineage-tracing data suggested that EEs can be made via three distinct routes (Figure 1): 1) Upon ISC division, one daughter retains ISC fate and one daughter differentiates to EE fate, 2) ISC division can produce 2 EE cells, thereby ending that stem cell lineage and 3) ISCs are likely capable of direct differentiation into EE fate [8], a finding that has recently been described in mammalian ISCs in culture upon induced quiescence [12]. Multiple studies now support the notion of a specific EE precursor cell [8,9,13,14,15]. However, whether this precursor is an ISC primed to produce EE cells [8,13,15,16], or a specialized EE precursor (EE Mother Cell, EMC) expressing defined markers distinct from the ISC and having limited division potential [14], has not yet been resolved. Importantly, additional cell fate markers, further lineage tracing strategies, and long-term live imaging will be required to resolve the exact lineage(s) producing the EE cell as well as how ISC plasticity is facilitated. Symmetric cell fate acquisition

Evidence suggests that, in addition to asymmetric divisions, ISCs can divide symmetrically with respect to daughter fate, resulting both in symmetric renewing divisions, generating 2 ISCs, or symmetric differentiation divisions in which the ISC is lost as both daughters Current Opinion in Cell Biology 2017, 48:97–105

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Figure 1

Future studies will be important to better understand how the ratio between symmetric and asymmetric divisions is regulated. Asymmetric cell fate acquisition

Notch signaling is necessary and sufficient for EC cell fate determination and must be kept off or low in ISCs and EE cells [3,4], a process that is only partly understood. Differential Notch activation between stem cell daughters was proposed to rely on Par complex-dependent intrinsic polarity that is thought to enhance Notch signaling. In this model, basement membrane adhesion via Integrins form an extrinsic cue to bias spindle orientation and Par complex control of Notch cell fate decisions [22]. How the Par complex might enhance binary Notch activation in the daughter cells was not clear in this model. However, follow-up studies showed that down-regulation of the basement membrane components Perlecan or b-integrin resulted in a loss of ISCs [23,24,25] instead of their accumulation as previously observed [22]. Another proposed asymmetrically segregating factor is Sara endosomes having enriched inheritance in the more apical ISC daughter cell and thought to help activate the Notch pathway in one of the daughter cells [26]. Furthermore, studies using overexpressed Dpp-GFP suggested that the basal secretion of BMP ligands results in a gradient of activation, inhibiting Notch signaling in the basal-most stem cell daughter cell through a yet to be identified mechanism [27]. Transcriptional control of the ISC

Schematic view of the Drosophila intestinal lineage. While intestinal stem cells are multipotent, recent evidence points toward the idea that EE and EC are formed from different intermediate precursor cells (EBs). High Notch signaling activation in the EB is required for them to differentiate into EC while low or no Notch activation would lead to EE precursors and EE formation. Evidence supports different paths to EE fate: an ISC producing an EE could 1) divide asymmetrically to produce one precursor that will eventually differentiate into an EE, 2) divide symmetrically to produce two EEs or 3) differentiate directly, without division to form an EE. EE fate acquisition requires Numb, an inhibitor of Notch signaling [11].

terminally differentiate [17]. Symmetric self-renewing divisions are less frequent during homeostasis though increase dramatically upon re-feeding after starvation in response to local secretion of an insulin-like peptide [17,18,19]. The RNA-binding protein Lin-28 has recently been shown to enhance Insulin receptor (InR) mRNA levels in the ISC [20]. InR signaling, in turn, leads to a decrease in miR-305 thereby increasing levels of a target mRNA, Hairless, implicated in silencing Notch transcriptional targets to promote symmetric self-renewal [21]. Current Opinion in Cell Biology 2017, 48:97–105

While additional mechanisms impacting de choice between asymmetric and symmetric fate of ISC daughters are likely, studies of the regulation of the transcriptional programs of ISCs and EBs fate are beginning to unveil important factors controlling ISC and EB cell identity and lineage progression. These factors include Daughterless, GATAe, escargot, Charlatan among other regionally expressed transcription factors [6,28,29,30,31]. In addition, more general chromatin-remodelling factors such as the histone deubiquitinase Scrawny, the Histone acetyl transferase ATAC2, and the SWI/SNF factor, Osa, also are required for proper stem cell function [32,33,34]. Understanding the epigenetic profile of each cell type and how it is modified during differentiation will be active areas for future research.

Local signals regulating ISC proliferative activity A brief overview of networks regulating ISC proliferation

The intestine undergoes slow ISC-mediated replacement of differentiated cells during normal homeostasis, though ISCs are not absolutely required for intestine function and normal longevity [35,36,37]. However, after tissue damage, ISCs become essential for viability and have a dramatically increased proliferation rate [37,38,39,40]. Modulation of ISC proliferation has been extensively www.sciencedirect.com

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reviewed elsewhere and relies on various signaling pathways among which Jak-Stat, EGFR, Dpp, Wingless (Wg), Hippo, Hh, Jnk and InR, and see for review [5,41,42]. New insight has begun to reveal how different ISC proliferation signals are integrated within the ISC. The use of an in vivo Ca2+ sensor demonstrated that EGFR, Jak-Stat, InR and Jnk signaling stimulate proliferation via cytosolic Ca2+ oscillations [43]. The proto-oncogene Myc is another central downstream factor important for optimal ISC proliferation after damage integrating at the transcriptional level many different signaling pathways in the ISC including Jak-Stat, EGFR, Wg and Hippo [44,45,46]. In addition, the transcription factor Sox21a has been suggested to act both in the ISC and EB cells to limit ISC proliferation in normal conditions and promote ISC division after stress [9,47,48]. Finally, another major regulator of the gut and its proliferative status is the sex of the fly, with the sex determination pathway required intrinsically in the ISCs and likely the ECs to drive sex-specific differences between males and females impacting intestine size, stem cell proliferation rates, responses to stress, and structure during aging [49,50]. Signals coming from the intestinal epithelium and surrounding visceral muscles

The epithelial cells and the surrounding visceral muscles (VM) are probably the most important and most direct sources of ligands of the signaling pathways controlling ISC proliferation (Figure 2). This is the case for (1) Wg ligand produced by the VM and the progenitor cells [44,51,52], (2) Jak-Stat ligands Upd1 expressed by the ISC/EBs [53] and the VM [54], while Upd2 and Upd3 are

expressed only by the epithelial cells (ECs/ISC/EBs and ECs respectively) [53,55] and (3) EGFR ligands, Vein being secreted by the VM, while Keren and Spitz are produced by the ECs and the EBs, respectively [56,57]. Signals from the trachea?

Tracheal cells, which associate closely with the intestine to deliver oxygen to the epithelial and surrounding tissues, have been shown to produce the Bmp ligand Dpp [58,59,60] (Figure 2). The exact role of Dpp from the trachea however remains controversial as two studies have opposing data on whether disruption of Dpp in trachea limits ISC proliferation [58] or not [59]. Additional sources of Dpp have been proposed: the ECs [27,61], the VM [59,60] and the haemocytes [62]. In addition, a second Bmp ligand, Gbb, is expressed in the ECs [27,60]. Bmp signaling also is required for regional identity specification of the middle midgut [59,61]. Further studies will be necessary to decipher additional signals that the trachea may send to the gut and the surrounding VM. These data suggest that despite not being localized in a physical structure such as the crypt of the mammalian intestine, neighboring cells of the epithelium (EBs, ECs, EEs) as well as surrounding tissues such the VM, and the trachea participate in the elaboration of a niche-like microenvironment controlling ISC activity.

Long-range signals regulating ISC proliferative activity In addition to short-range signals from epithelial cells or adjacent tissues, more recent studies demonstrate the

Figure 2

Overview of the tissues communications regulating ISC activity. In physiological conditions of homeostasis, or after damage of the tissue, the ligands of the signaling pathways controlling ISC proliferation are often produced locally by epithelial cells and by surrounding tissues such as the trachea (Dpp) and the visceral muscles (Upd1, Vein, Wg, Dilp3). Long-range signals participate also to this regulation of ISC activity with ligands produced by the brain (Dilps), the neurons (Upds), the corpus allatum (JH) and the haemocytes after injury or infection (Upd3, Dpp). The intestine is not only an integrator of signals but is also able to send signals toward the brain such as ImpL2 to reduce insulin pathway activity, or toward the fat body through Hh released by the ECs and Activin-b by the EEs to regulate metabolism. www.sciencedirect.com

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importance of long-range signals capable of remotely controlling ISC behaviour. Indeed, multiple sources of the ligands of signaling pathways regulating ISC activity often have local as well as more distant tissue sources (Figure 2). Signals from the brain

Both gut epithelial cells and the VM are directly innervated by neurons originating from the central and peripheral nervous system that control dietary and metabolic functions [63]. Damage to these neurons can produce a proliferative response of the ISCs, which was suggested to rely on neuronal Hedgehog (Hh) signaling mediated by EC cells-secreted Hh [64]. Hh signaling in EB cells and VM was also detected and promotes ISC proliferation after DSS-induced injury [65,66]. Insulin-like peptides (Dilps) produced in the brain may also contribute to control ISC proliferation as their ablation resembles knock-down of InR in the ISC [39]. In addition, Dilp3 is produced locally in the VM of a specific region of the posterior midgut and activates ISC division upon refeeding, thereby linking cell division kinetics to food availability [18,39]. Signals from the corpus allatum, an endocrine organ

The intestinal epithelium also receives input regarding mating behaviour of the fly: upon copulation, female flies have an increase in Juvenile hormone (JH) produced in the corpus allatum in response to male-derived Sex Peptide. JH coordinates an array of metabolic changes in preparation for egg production including promoting an increased number of ISC divisions and an expansion of the intestine [67]. Whether, symmetric divisions are preferentially increased similar to Dilp3-mediated ISC division upon re-feeding after starvation [18], has not been investigated.

the abdomen, leads to the degeneration of fat body, ovaries and muscles, reminiscent to mammalian organ wasting often associated with cancer or chronic pathologies. The degenerate effect is due to the secretion of an antagonist of insulin pathway (Impl2) by the gut reducing systemically insulin pathway activity [70,71]. The gut ECs are also a major site of Hedgehog production, which appears to act long-range to impact metabolism in the fat body (at least in the larval gut) as well as locally on neurons innervating the gut, and within EBs to control proliferation [65,66,72]. Studies in the larval gut recently demonstrated that upon a high sugar diet, EE cells produce Activin-b (Act-b) received by the fat-body to activate AKH/glucagon pathway for glycemic control [73]. Further studies will determine if additional gut-derived systemic signals exist. Together these works show how organ — organ communication as well as intra-organ communication are essential to the regulation of adult SC activity during physiological homeostasis as well as in regenerative or pathologic conditions.

Aging and mechanisms to cope with cellular damage Alteration of gut structure and bacterial populations during aging

ISCs can be remotely activated to divide in response to a body puncture wound [68]. Recent work has further elucidated how this response is coordinated: upon septic or aseptic injury, haemocytes secrete Upd3 leading to JakStat activation in progenitor cells and VM cells thereby stimulating ISC proliferation [69]. Upon oral infection, haemocytes were proposed to be recruited to the midgut, secrete Dpp, and be necessary for a regenerative response [62]. A separate study, however, did not detect an induced recruitment of haemocytes and found that they were not essential for ISC proliferative divisions upon bacterial oral infection [69]. How haemocytes detect septic injury remains unclear, though may be mediated by generation of reactive oxygen species [68].

During aging, in humans and flies alike, adult tissues experience changes in structure and function accompanied by alteration in stem cell self-renewal and differentiation properties [1]. In the midgut upon aging, the middle, Copper cell region of the midgut undergoes alteration of cell identity with a loss of acid producing cells [74] which, in turn, provokes dysbiosis of commensal bacteria. Concomitantly, an alteration of gut epithelial integrity and permeabilization of the intestinal barrier occur due to a loss of tricellular junctions between EC during aging [75,76,77,78,79,80,81,82]. Following dysbiosis and intestinal barrier break-down an age-dependent increase in Jnk and Jak-Stat signaling lead to increased ISC proliferation, resulting in a piling up of undifferentiated EBs, potentially further disrupting the intestinal barrier (Figure 3). Interestingly, lifespan can be extended by limiting commensal bacteria or decreasing ISC proliferation, at least in female flies [50,78,81,83,84]. An age-associated increase in the growth factor PVF2 (a fly ortholog of Platelet Derived Growth Factor and Vascular Endothelial Frowth Factors) also stimulates ISC proliferation and likely acts downstream of the p38 stress signaling pathway [83,85]. Whether p38 signaling may be activated upon epithelial barrier defects is currently not known.

Signals sent from the gut

DNA damage and genome modification during aging

Recent studies highlight how the gut can be the transmitter of systemic signals. Yki-induced midgut overproliferation, or the transplantation of tumorous tissues into

In addition to important changes to epithelial integrity and gut microbiome in the aging gut, there is also an increase in marks of damaged proteins and DNA,

Signals from haemocytes

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Figure 3

Age related loss of intestinal homeostasis. Upon aging the Drosophila gut structure and function gradually declines. Dysbiosis of the commensal bacteria is associated with loss of the intestinal epithelium integrity due to alteration of tricellular junctions (TCJ). This further results in permeabilization of the intestinal barrier, ISC overproliferation and dysplasia formation by accumulation of undifferentiated EBs. During aging, ISCs also accumulates spontaneous mutations as illustrated by neoplasia formation in 10% of aged wild-type males due to inactivation of the X-linked gene Notch.

including marks of DNA double-strand break and reactive oxygen-induced damage [86,87,88]. Moreover, recent findings have demonstrated that DNA damage is not all correctly repaired and has important consequences on the aging genome, resulting in frequently arising spontaneous mutations [89]. Indeed, loss-ofheterozygosity as well as deletions and chromosome www.sciencedirect.com

rearrangements occur frequently during aging. Consequently, in roughly 10% of aged male flies (having a single X chromosome), spontaneous inactivation of the Notch tumor suppressor gene (localized on the X chromosome) generates neoplasia formation, representing approximately 1 Notch mutant ISC in 10,000 ISCs [89] (Figure 3). Of note, the intestine has robust Current Opinion in Cell Biology 2017, 48:97–105

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mechanisms of cell competition that can eliminate cells that are less fit due to ribosomal mutations [90]. Whether these mechanisms also help eliminate genetically unfit mutant cells is currently unclear. However, once a Notch mutant ISC gives rise to a small mass of cells, cell competition mechanisms and induction of a surrounding “niche” actually further drive its growth process [91,92]. Interestingly, recent findings have demonstrated that wild-type ISCs, which are usually relatively quiescent, do not respond as other cells to induction of the proapototic genes reaper and p53, instead undergoing autophagy-mediated death upon loss of lipolysis [35,93,94]. Only in rapidly dividing mutant contexts such as upon Notch inhibition or Raf gain-of-function, can reaper promote apoptosis [93,94]. It is possible that upon induction of proliferation, DNA lesions present in quiescent cells in or G1-arrested cells will be converted into DNA double-strand breaks during S phase, thereby activating checkpoints that facilitate apoptosis-mediated cell death. Of note, apoptosis-mediated death of ISCs can occur in response to cell competition cues [90]. Therefore, future research will be needed to understand what types of damage can and cannot trigger cell elimination and the precise roles of autophagy and apoptosis in quiescent and actively dividing stem cells. Ultimately, knowing how cells evade detection and elimination, resulting in mutant and abnormal cells during aging, will provide strategies to promote healthy aging.

Acknowledgements Apologies to colleagues whose work was not cited due to space constraints. We would like to thank members of the Bardin lab for comments on the manuscript. The Bardin lab is supported by the CNRS, Inserm, Institute Curie and Project grants from the Fondation ARC, ANR (SoMuSeqSTEM), by “Labelisation FRM”, and by the “Fondation Schlumberger pour l’Education et la Recherche”.

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This study presents evidences that macrophage-like haemocytes are recruited to the intestine specifically after injury and produce Dpp ligand promoting increased ISC proliferation during the regenerative stage followed by repression of their proliferation during recovery. 63. Cognigni P, Bailey AP, Miguel-Aliaga I: Enteric neurons and systemic signals couple nutritional and reproductive status with intestinal homeostasis. Cell Metabol 2011, 13:92-104. 64. Han H, Pan C, Liu C, Lv X, Yang X, Xiong Y, Lu Y, Wu W, Han J, Zhou Z et al.: Gut-neuron interaction via Hh signaling regulates intestinal progenitor cell differentiation in Drosophila. Cell Discov 2015, 1:15006-15013. 65. Li Z, Guo Y, Han L, Zhang Y, Shi L, Huang X, Lin X: Debramediated Ci degradation controls tissue homeostasis in Drosophila adult midgut. Stem Cell Reports 2014, 2:135-144. 66. Tian A, Shi Q, Jiang A, Li S, Wang B, Jiang J: Injury-stimulated Hedgehog signaling promotes regenerative proliferation of Drosophila intestinal stem cells. J Cell Biol 2015, 208:807-819. 67. Reiff T, Jacobson J, Cognigni P, Antonello Z, Ballesta E, Tan KJ,  JY Y, Dominguez M, Miguel-Aliaga I: Endocrine remodelling of the adult intestine sustains reproduction in Drosophila. eLife 2015 http://dx.doi.org/10.7554/eLife.06930.001. This study shows how female flies ensure sufficient nutrients for their progeny by releasing Juvenile Hormone (JH) after mating. JH induces ISC proliferation, leading to an increase of the numbers of cells in the midgut and an increase of its size. 68. Takeishi A, Kuranaga E, Tonoki A, Misaki K, Yonemura S, Kanuka H, Miura M: Homeostatic epithelial renewal in the gut is required for dampening a fatal systemic wound response in Drosophila. Cell Reports 2013, 3:919-930. 69. Chakrabarti S, Dudzic JP, Li X, Collas EJ, Boquete J-P, Lemaitre B:  Remote control of intestinal stem cell activity by haemocytes in Drosophila. PLoS Genet 2016, 12:e1006089-e1006124. In this work, the authors show how after septic injury, the haemocytes secrete Upd3 leading to Jak-STAT activation in the ISC and thereby stimulating their proliferation and gut production of microbial peptides. This control of intestine renewal by haemocytes is proposed to be part of a survival program essential to Drosophila resistance in response to injury while being dispensable for response to oral infection. 70. Kwon Y, Song W, Droujinine IA, Hu Y, Asara JM, Perrimon N:  Systemic organ wasting induced by localized expression of the secreted insulin/IGF antagonist ImpL2. Dev Cell 2015, 33:36-46. Using overexpression of Yorkie in the ISC, this study investigated how an overproliferating tissue can lead to organ wasting phenotypes in vivo. They unveiled the role in this phenomenon of insulin antagonist (ImpL2) secretion by the intestine which represses systemically the insulin like pathway while the overproliferating gut is able to escape organ wasting by local activation of insulin signaling. 71. Figueroa-Clarevega A, Bilder D: Malignant drosophila tumors  interrupt insulin signaling to induce cachexia-like wasting. Dev Cell 2015, 33:47-55. Figueroa-Clareverga and Bilder show that transplantation of imaginal disc tumors into the abdomen of wild-type flies leads to organ wasting phenotypes downstream of ImpL2 secreted by the overproliferating tissue and systemic inhibition of insulin pathways. 72. Rodenfels J, Lavrynenko O, Ayciriex S, Sampaio JL, Carvalho M, Shevchenko A, Eaton S: Production of systemically circulating Hedgehog by the intestine couples nutrition to growth and development. Genes Dev 2014, 28:2636-2651. 73. Song W, Cheng D, Hong S, Sappe B, Hu Y, Wei N, Zhu C, O’Connor MB, Pissios P, Perrimon N: Midgut-derived activin regulates glucagon-like action in the fat body and glycemic control. Cell Metabol 2017, 25:386-399. 74. Li H, Jasper H: Gastrointestinal stem cells in health and disease: from flies to humans. Dis Model Mech 2016, 9:487-499. 75. Ren C, Webster P, Finkel SE, Tower J: Increased internal and external bacterial load during Drosophila aging without lifespan trade-off. Cell Metabol 2007, 6:144-152. 76. Rera M, Bahadorani S, Cho J, Koehler CL, Ulgherait M, Hur JH, Ansari WS, Lo T Jr, Jones DL, Walker DW: Modulation of longevity and tissue homeostasis by the Drosophila PGC-1 homolog. Cell Metabol 2011, 14:623-634. Current Opinion in Cell Biology 2017, 48:97–105

77. Rera M, Clark RI, Walker DW: Intestinal barrier dysfunction links metabolic and inflammatory markers of aging to death in Drosophila. Proc Natl Acad Sci USA 2012, 109:21528-21533. 78. Clark RI, Salazar A, Yamada R, Fitz-Gibbon S, Morselli M, Alcaraz J, Rana A, Rera M, Pellegrini M, Ja WW et al.: Distinct shifts in microbiota composition during Drosophila aging impair intestinal function and drive mortality. Cell Reports 2015, 12:1656-1667. 79. Buchon N, Broderick NA, Chakrabarti S, Lemaitre B: Invasive and indigenous microbiota impact intestinal stem cell activity through multiple pathways in Drosophila. Genes Dev 2009, 23:2333-2344. 80. Broderick NA, Buchon N, Lemaitre B: Microbiota-induced changes in drosophila melanogaster host gene expression and gut morphology. mBio 2014, 5:e01117-e1214. 81. Guo L, Karpac J, Tran SL, Jasper H: PGRP-SC2 promotes gut immune homeostasis to limit commensal dysbiosis and extend lifespan. Cell 2014, 156:109-122. 82. Resnik-Docampo M, Koehler CL, Clark RI, Schinaman JM,  Sauer V, Wong DM, Lewis S, D’Alterio C, Walker DW, Jones DL: Tricellular junctions regulate intestinal stem cell behaviour to maintain homeostasis. Nat Cell Biol 2016, 19:52-59. This study reports how altered organization of tricellular junctions between cells of the Drosophila intestinal epithelium leads to ISC overproliferation and misdifferentiation. A gradual loss of TCJ is observed upon aging and could be a factor important in age-related decline of the tissue. 83. Choi N-H, Kim J-G, Yang D-J, Kim Y-S, Yoo M-A: Age-related changes in Drosophila midgut are associated with PVF2, a PDGF/VEGF-like growth factor. Aging Cell 2008, 7:318-334. 84. Biteau B, Karpac J, Supoyo S, Degennaro M, Lehmann R, Jasper H: Lifespan extension by preserving proliferative homeostasis in Drosophila. PLoS Genet 2010, 6:e1001159. 85. Park J-S, Kim Y-S, Kim J-G, Lee S-H, Park S-Y, Yamaguchi M, Yoo M-A: Regulation of the Drosophila p38b gene by transcription factor DREF in the adult midgut. BBA - Gene Regulat Mech 2010, 1799:510-519. 86. Bufalino MR, DeVeale B, van der Kooy D: The asymmetric segregation of damaged proteins is stem cell-type dependent. J Cell Biol 2013, 201:523-530. 87. Park J-S, Lee S-H, Na H-J, Pyo J-H, Kim Y-S, Yoo M-A: Age- and oxidative stress-induced DNA damage in Drosophila intestinal stem cells as marked by Gamma-H2AX. EXG 2012, 47:401-405. 88. Park JS, Na HJ, Pyo JH, Jeon HJ, Kim YS, Yoo M-A: Requirement of ATR for maintenance of intestinal stem cells in aging Drosophila. Aging (Albany NY) 2015, 7:307-318. 89. Siudeja K, Nassari S, Gervais L, Skorski P, Lameiras S, Stolfa D,  Zande M, Bernard V, Frio TR, Bardin AJ: Frequent somatic mutation in adult intestinal stem cells drives neoplasia and genetic mosaicism during aging. Cell Stem Cell 2015, 17:663-674. The authors show that frequent spontaneous mutations are acquired by ISC during aging. These observed mutation events correspond to loss of heterozygosity via mitotic recombination or to spontaneous deletions and chromosome rearrangements. Consequently, in 10% of aged male flies, inactivation of the single copy of the Notch gene located on the X chromosome leads to the formation of large neoplastic growths. 90. Kolahgar G, Suijkerbuijk SJE, Kucinski I, Poirier EZ, Mansour S,  Simons BD, Piddini E: Cell competition modifies adult stem cell and tissue population dynamics in a JAK-STAT-dependent manner. Dev Cell 2015, 34:297-309. Using lineage tracing experiments and physical modeling, this study shows the existence of cell competition in the fly intestine allowing the elimination of unfit cells by healthy tissue and highlights the role of JakSTAT pathway in this process, which induces winner ISC proliferation and self-renewal. 91. Suijkerbuijk SJE, Kolahgar G, Kucinski I, Piddini E: Cell  competition drives the growth of intestinal adenomas in Drosophila. Curr Biol 2016, 26:428-438. www.sciencedirect.com

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Using APC mutant clones, the authors show that overproliferating adenoma cells compete and kill surrounding cells and how preventing cell competition appears to be able to prevent mutant tissue growth. 92. Patel PH, Dutta D, Edgar BA: Niche appropriation by Drosophila  intestinal stem cell tumours. Nat Cell Biol 2015, 17:1182-1192. Using Notch defective clones to produce tumor-like ISC, Patel et al., demonstrate the role of surrounding niche signals induced non-autonomously by the tumor cells to drive further growth.

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93. Singh SR, Zeng X, Zhao J, Liu Y, Hou G, Liu H, Hou SX: The lipolysis pathway sustains normal and transformed stem cells in adult Drosophila. Nature 2016, 538:109-113. 94. Ma M, Zhao H, Zhao H, Binari R, Perrimon N, Li Z: Wildtype adult stem cells, unlike tumor cells, are resistant to cellular damages in Drosophila. Dev Biol 2016, 411:207-216.

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