The production of offspring is energetically costly and relies on incompletely understood mechanisms that generate a positive energy balance. In mothers of many species, changes in key energy-associated internal organs are common yet poorly characterised functionally and mechanistically. In this study, we show that, in adult Drosophila females, the midgut is dramatically remodelled to enhance reproductive output. In contrast to extant models, organ remodelling does not occur in response to increased nutrient intake and/or offspring demands, but rather precedes them. With spatially and temporally directed manipulations, we identify juvenile hormone (JH) as an anticipatory endocrine signal released after mating. Acting through intestinal bHLH-PAS domain proteins Methoprene-tolerant (Met) and Germ cell-expressed (Gce), JH signals directly to intestinal progenitors to yield a larger organ, and adjusts gene expression and sterol regulatory element-binding protein (SREBP) activity in enterocytes to support increased lipid metabolism. Our findings identify a metabolically significant paradigm of adult somatic organ remodelling linking hormonal signals, epithelial plasticity, and reproductive output.DOI: http://dx.doi.org/10.7554/eLife.06930.001

The intestinal epithelium is remarkably robust despite perturbations and demand uncertainty. Here, we investigate the basis of such robustness using novel tracing methods that allow simultaneously capturing the dynamics of stem and committed progenitor cells (called enteroblasts) and intestinal cell turnover with spatiotemporal resolution. We found that intestinal stem cells (ISCs) divide “ahead” of demand during Drosophila midgut homeostasis. Their newborn enteroblasts, on the other hand, take on a highly polarized shape, acquire invasive properties and motility. They extend long membrane protrusions that make cell–cell contact with mature cells, while exercising a capacity to delay their final differentiation until a local demand materializes. This cellular plasticity is mechanistically linked to the epithelial–mesenchymal transition (EMT) programme mediated by escargot, a snail family gene. Activation of the conserved microRNA miR-8/miR-200 in “pausing” enteroblasts in response to a local cell loss promotes timely terminal differentiation via a reverse MET by antagonizing escargot. Our findings unveil that robust intestinal renewal relies on hitherto unrecognized plasticity in enteroblasts and reveal their active role in sensing and/or responding to local demand.

The transcription factor Gata3 is essential for the development of sympathetic neurons and adrenal chromaffin cells. As Gata3 expression is maintained up to the adult stage, we addressed its function in differentiated sympathoadrenal cells at embryonic and adult stages by conditional Gata3 elimination. Inactivation of Gata3 in embryonic DBH-expressing neurons elicits a strong reduction in neuron numbers due to apoptotic cell death and reduced proliferation. No selective effect on noradrenergic gene expression (TH and DBH) was observed. Interestingly, Gata3 elimination in DBH-expressing neurons of adult animals also results in a virtually complete loss of sympathetic neurons. In the Gata3-deficient population, the expression of anti-apoptotic genes (Bcl-2, Bcl-x L , and NFB) is diminished, whereas the expression of pro-apoptotic genes (Bik, Bok, and Bmf) was increased. The expression of noradrenergic genes (TH and DBH) is not affected. These results demonstrate that Gata3 is continuously required for maintaining survival but not differentiation in the sympathetic neuron lineage up to mature neurons of adult animals.

SUMMARYNeuroblastoma (NB) is the most common extracranial solid tumor in childhood and arises from cells of the developing sympathoadrenergic lineage. Activating mutations in the gene encoding the ALK tyrosine kinase receptor predispose for NB. Here, we focus on the normal function of Alk signaling in the control of sympathetic neuron proliferation, as well as on the effects of mutant ALK. Forced expression of wild-type ALK and NB-related constitutively active ALK mutants in cultures of proliferating immature sympathetic neurons results in a strong proliferation increase, whereas Alk knockdown and pharmacological inhibition of Alk activity decrease proliferation. Alk activation upregulates NMyc and trkB and maintains Alk expression by an autoregulatory mechanism involving Hand2. The Alk-ligand Midkine (Mk) is expressed in immature sympathetic neurons and in vivo inhibition of Alk signaling by virus-mediated shRNA knockdown of Alk and Mk leads to strongly reduced sympathetic neuron proliferation. Taken together, these results demonstrate that the extent and timing of sympathetic neurogenesis is controlled by Mk/Alk signaling. The predisposition for NB caused by activating ALK mutations may thus be explained by aberrations of normal neurogenesis, i.e. elevated and sustained Alk signaling and increased NMyc expression. Activation of Alk signaling, in particular overexpression of ALK wt and NB ALK mutants resulted in the upregulation of NMyc and trkB, which may contribute to continued proliferation and NB predisposition. MATERIALS AND METHODS Expression plasmidsPcDNA3.1 expression plasmids for human ALK wt , ALK F1174L and ALK R1275Q, and pCAGGS expression plasmids for Hand2, Phox2b wt and Phox2b K155X have been described previously (Janoueix-Lerosey et al., 2008;Reiff et al., 2010). shRNA constructsThe shRNA against Gallus gallus Alk and Mk were designed using BLOCK-iT RNAi Designer (Invitrogen, Karlsruhe, Germany) and target the following sequences: shAlk, 5Ј-AAU GGU UUC UCU CUA UGU CCA ACU C-3Ј; shMk, 5Ј-GAG CUG ACU GCA AGU ACA AGU UUG A-3Ј. Scrambled shRNAs (sc-shRNA, Invitrogen, Karlsruhe, Germany) served as controls. In situ hybridizationNon-radioactive in situ hybridization on cryosections and preparation of digoxigenin labeled probes for chick Phox2b was carried out as described previously (Ernsberger et al., 1997;Stanke et al., 1999). qPCR analysisEqual amounts of RNA were used to synthesize cDNA with Oligo(dT)-primers and Superscript-III-reverse-transcriptase according to manufacturer's instructions (Invitrogen, Karlsruhe, Germany). The PCR was carried out using Abgene's Absolute Blue SYBR-Green qPCR Mix (Abgene, Epsom, UK) in a Stratagene Mx3000p Light Cycler (Stratagene, Waldbronn, Germany). All primers (MWG Biotech AG, Ebersberg, Germany) were designed to anneal optimally at 58°C with PerlPrimer (Marshall, 2004) (95°C for 30 seconds, 58°C for 30 seconds, 72°C for 30 seconds, repeated for 50 cycles and a subsequent dissociation curve). The primer pairs (see Table S1 in the supplementary material) were ...

Developmental studies revealed fundamental principles on how organ size and function is achieved, but less is known about organ adaptation to new physiological demands. In fruit flies, juvenile hormone (JH) induces intestinal stem cell (ISC) driven absorptive epithelial expansion balancing energy uptake with increased energy demands of pregnancy. Here, we show 20-Hydroxy-Ecdysone (20HE)-signaling controlling organ homeostasis with physiological and pathological implications. Upon mating, 20HE titer in ovaries and hemolymph are increased and act on nearby midgut progenitors inducing Ecdysone-induced-protein-75B (Eip75B). Strikingly, the PPARγ-homologue Eip75B drives ISC daughter cells towards absorptive enterocyte lineage ensuring epithelial growth. To our knowledge, this is the first time a systemic hormone is shown to direct local stem cell fate decisions. Given the protective, but mechanistically unclear role of steroid hormones in female colorectal cancer patients, our findings suggest a tumor-suppressive role for steroidal signaling by promoting postmitotic fate when local signaling is deteriorated.

The regenerative activity of adult stem cells carries a risk of cancer, particularly in highly renewable tissues. Members of the family of inhibitor of apoptosis proteins (IAPs) inhibit caspases and cell death, and are often deregulated in adult cancers; however, their roles in normal adult tissue homeostasis are unclear. Here, we show that regulation of the number of enterocyte‐committed progenitor (enteroblast) cells in the adult Drosophila involves a caspase‐mediated physiological apoptosis, which adaptively eliminates excess enteroblast cells produced by intestinal stem cells (ISCs) and, when blocked, can also lead to tumorigenesis. Importantly, we found that Diap1 is expressed by enteroblast cells and that loss and gain of Diap1 led to changes in enteroblast numbers. We also found that antagonistic interplay between Notch and EGFR signalling governs enteroblast life/death decisions via the Klumpfuss/WT1 and Lozenge/RUNX transcription regulators, which also regulate enteroblast differentiation and cell fate plasticity. These data provide new insights into how caspases drive adult tissue renewal and protect against the formation of tumours.

Neuroblastoma is a pediatric tumor that is thought to arise from autonomic precursors in the neural crest. Mutations in the PHOX2B gene have been observed in familial and sporadic forms of neuroblastoma and represent the first defined genetic predisposition for neuroblastoma. Here, we address the mechanisms that may underlie this predisposition, comparing the function of wild-type and mutant Phox2b proteins ectopically expressed in proliferating, embryonic sympathetic neurons. Phox2b displays a strong antiproliferative effect, which is lost in all Phox2b neuroblastoma variants analyzed. In contrast, an increase in sympathetic neuron proliferation is elicited by Phox2b variants with mutations in the homeodomain when endogenous Phox2b levels are lowered by siRNA-mediated knockdown to mimic the situation of heterozygous PHOX2B mutations in neuroblastoma. The increased proliferation is blocked by Hand2 knockdown and the antiproliferative Phox2b effects are rescued by Hand2 overexpression, implying Hand2 in Phox2b-mediated proliferation control. A Phox2b variant with a nonsense mutation in the homeodomain elicits, in addition, a decreased expression of characteristic marker genes. Together, these results suggest that PHOX2B mutations predispose to neuroblastoma by increasing proliferation and promoting dedifferentiation of cells in the sympathoadrenergic lineage.

Neural crest stem cells (NCSCs) give rise to the neurons and glia of the peripheral nervous system (PNS). NCSC-like cells can be isolated from multiple peripheral organs and maintained in neurosphere culture. Combining in vitro culture and transplantation, we show that expanded embryonic NCSC-like cells lose PNS traits and are reprogrammed to generate CNS cell types. When transplanted into the embryonic or adult mouse CNS, they differentiate predominantly into cells of the oligodendrocyte lineage without any signs of tumor formation. NCSC-derived oligodendrocytes generate CNS myelin and contribute to the repair of the myelin deficiency in shiverer mice. These results demonstrate a reprogramming of PNS progenitors to CNS fates without genetic modification and imply that PNS cells could be a potential source for cell-based CNS therapy.