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Cell Stem Cell
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MT-Nanotubes: Lifelines for Stem Cells

  • Marc Amoyel
    Affiliations
    Department of Biochemistry and Molecular Pharmacology, The Helen L. and Martin S. Kimmel Center for Stem Cell Biology, New York University School of Medicine, 550 First Avenue, MSB 497B, New York, NY 10016, USA
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  • Erika A. Bach
    Correspondence
    Corresponding author
    Affiliations
    Department of Biochemistry and Molecular Pharmacology, The Helen L. and Martin S. Kimmel Center for Stem Cell Biology, New York University School of Medicine, 550 First Avenue, MSB 497B, New York, NY 10016, USA
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      Niche cells produce secreted factors that promote the self-renewal of stem cells in their immediate proximity, but how signaling is restricted to stem cells is not well understood.
      • Inaba M.
      • Buszczak M.
      • Yamashita Y.M.
      report that microtubule (MT) structures called MT-nanotubes control activation of the primary self-renewal pathway in Drosophila testes.

      Main Text

      A longstanding question in stem cell biology is how diffusible self-renewal signals produced by a niche are restricted to stem cells and not to other cells in the vicinity of this niche. Several niche-derived self-renewal factors, including Bone Morphogenetic Proteins (BMPs), are also morphogens, raising the additional issue of how long-range factors can act as short-range, binary on-off signals in niche-stem cell interactions. Some strategies to explain the restricted action of self-renewal signals have been proposed, including specific subcellular localization of self-renewal signal transduction (
      • Michel M.
      • Raabe I.
      • Kupinski A.P.
      • Pérez-Palencia R.
      • Bökel C.
      ). In a recent study in Nature, Yamashita and colleagues offer an additional mechanism. They characterize signal-bearing MT-based structures emanating from germline stem cells (GSCs) toward niche cells in the Drosophila testis that explain how only GSCs and not their differentiating daughter cells can access self-renewal signals from the niche (
      • Inaba M.
      • Buszczak M.
      • Yamashita Y.M.
      ).
      The Drosophila testis is one of the best characterized niche-stem cell systems, as both the niche and stem cells are anatomically and molecularly well-defined, self-renewal signals have been identified, and this tissue can be subjected to ex vivo live imaging (
      • Spradling A.
      • Fuller M.T.
      • Braun R.E.
      • Yoshida S.
      ). In the testis, the niche is composed of post-mitotic somatic cells termed hub cells. The niche maintains two stem cell populations, GSCs and somatic cyst stem cells (CySCs). Hub cells produce the BMPs Decapentaplegic (Dpp) and Glass bottom boat (Gbb), which are bona fide GSC self-renewal factors. More specifically, when GSCs are unable to activate BMP signaling, because they either lack the BMP receptor Thickveins (Tkv) or the R-Smad transcription factor Mothers against dpp (Mad), they cannot self-renew and are lost from the niche (
      • Kawase E.
      • Wong M.D.
      • Ding B.C.
      • Xie T.
      ,
      • Shivdasani A.A.
      • Ingham P.W.
      ). During mitosis, GSCs orient their mitotic spindle perpendicular to the niche. This alignment results in one GSC daughter cell being retained in the niche and hence preserving stem cell identity, while the other daughter cell (called a gonialblast) is displaced away from the niche and initiates differentiation (Figure 1). CySCs produce cyst cells, two of which ensheath one gonialblast. Transit-amplifying divisions of the gonialblast result in a 16-cell spermatogonial germ cyst that ultimately gives rise to spermatids.
      Figure thumbnail gr1
      Figure 1Schematic of the Apex of the Adult Drosophila Testis
      The interface between one hub cell and one GSC is boxed and enlarged at right. Hub cells secrete the BMP factor Dpp into the extracellular space. MT-based membrane protrusions, termed MT-nanotubes, emanate from GSCs and orient toward hub cells. Tkv, a BMP receptor, trafficks on these structures and becomes activated along the MT-nanotubes by secreted Dpp. This results in the phosphorylation, activation, and nuclear translocation of the Smad protein Mad (phosphoMad), which induces GSC self-renewal (circular arrow). Depletion of several IFT-B proteins from GSCs results in fewer and shorter nanotubes, with the consequence of less Tkv stimulation, reduced phosphoMad levels, and impaired self-renewal. By contrast, depletion of the kinesin Klp10A results in thicker nanotubes, augmented Tkv activation on MT-nanotubes, increased phosphoMad levels, and enhanced self-renewal.
      In the current study, Inaba et al. performed live imaging on unfixed Drosophila testes in which GFP-labeled α-tubulin was expressed in the germline lineage. This experiment revealed that MT-based structures, which they called MT-nanotubes, emanating from GSCs were oriented predominantly toward niche cells. MT-nanotubes were contained within a membrane bilayer and formed during interphase and S phase but disappeared during mitosis. These structures were sensitive to fixation, presumably explaining why they had not been previously observed. MT-nanotubes were still observed after treatment with the actin-depolymerizing drug cytochalasin B but were lost after treatment with the MT-depolymerizing drug colcemid, raising the possibility that MT-nanotubes were actually primary cilia. This hypothesis was disproved because, unlike primary cilia, MT-nanotubes were sensitive to fixation and were not necessarily associated with the basal body. Nevertheless, Inaba et al. found that MT-nanotubes depended on several factors found in primary cilia. They specifically demonstrated that germline depletion of Oseg2, Osm6, and Che-13, which are intraflagellar transport-B (IFT-B) components required for anterograde transport and assembly of the primary cilium, resulted in fewer and shorter nanotubes. By contrast, depletion of Klp10A, a member of the MT-depolymerizing kinesin family, significantly increased nanotube thickness.
      Having established the molecular characteristics of MT-nanotubes including positive and negative regulators, Inaba and colleagues next turned their attention to the function of these structures. They demonstrated that when the BMP receptor Tkv was expressed in the germline, it accumulated in puncta on MT-nanotubes that oriented toward the niche. Furthermore, they showed that Tkv proteins localized on MT-nanotubes adopted an activated state, suggesting that BMP signal transduction was initiated on MT-nanotubes. Indeed, BMP signaling in GSCs was reduced when MT-nanotubes were compromised and increased when MT-nanotubes were enlarged (Figure 1). Conversely, BMPs and their receptor were necessary and sufficient for MT-nanotube formation. This scenario leads to a model in which MT-nanotubes were necessary for BMP pathway activation yet paradoxically required BMPs in order to form in the first place. Even so, self-renewal was compromised in single GSCs deficient for IFT-B components, which is consistent with loss of BMP signaling in these mutant cells. It is noteworthy that lineage-wide depletion of MT-nanotube components did not result in global depletion of GSCs, which was the expected outcome of BMP loss (
      • Kawase E.
      • Wong M.D.
      • Ding B.C.
      • Xie T.
      ,
      • Shivdasani A.A.
      • Ingham P.W.
      ). It is possible that MT-nanotubes are a fidelity mechanism to ensure that BMP signaling is restricted to a GSC adhering to the niche but that a low level of BMP signaling still occurs in their absence. Importantly, GSCs also depend on CySCs as a source of BMPs (
      • Kawase E.
      • Wong M.D.
      • Ding B.C.
      • Xie T.
      ,
      • Leatherman J.L.
      • Dinardo S.
      ). Since no MT-nanotubes were observed oriented toward CySCs, this suggests that some BMP signaling in this tissue is MT-nanotube independent (
      • Inaba M.
      • Buszczak M.
      • Yamashita Y.M.
      ).
      Intriguingly, membrane extensions involved in intercellular signaling have been described in developing tissues across the animal kingdom, including in mice and flies (
      • Roy S.
      • Huang H.
      • Liu S.
      • Kornberg T.B.
      ,
      • Sanders T.A.
      • Llagostera E.
      • Barna M.
      ). Actin-based specialized filopodia shuttle the morphogens Dpp and Hedgehog (Hh) from the producing cells to the receiving cells located many cell diameters away, typically 40–150 μm. Ligands and activated receptors co-localize on these structures, similar to Dpp and Tkv on MT-nanotubes. Furthermore, cellular processes have been described in the GSC niche in C. elegans (
      • Byrd D.T.
      • Knobel K.
      • Affeldt K.
      • Crittenden S.L.
      • Kimble J.
      ). In this tissue, the niche cell called the distal tip cell (DTC) prevents the differentiation of GSCs located many cell diameters away. The DTC possesses a network of long extensions, presumably to deliver the membrane-bound self-renewal signal to distant GSCs. The common feature of these situations is that cellular projections are used to extend the range of signaling, unlike that described by Inaba et al., wherein MT-nanotubes serve to restrict BMP activation only at the niche-stem cell interface. It is notable that other insect species display extensions from GSCs toward neighboring niche cells, although it is not clear whether they are analogous to MT-nanotubes (
      • Schmidt E.D.
      • Dorn A.
      ). Nevertheless, GSC projections may represent a common feature of niche-stem cell communication in insects and other organisms. Future work should establish whether delivery of self-renewal factors by cellular projections is a broadly employed strategy and what determines whether stem cell niches utilize short- or long-range transport capabilities.

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