X
Forgot Password

If you have forgotten your password you can enter your email here and get a temporary password sent to your email.

GFP antibody

RRID:AB_305643

Antibody ID

AB_305643

Target Antigen

GFP antibody

Proper Citation

(Abcam Cat# ab6673, RRID:AB_305643)

Clonality

polyclonal antibody

Comments

validation status unknown, seller recommendations provided in 2012: ELISA; Immunohistochemistry - fixed; Immunohistochemistry - frozen; Immunohistochemistry; Western Blot; Immunoprecipitation; ELISA, IHC-Fr, IHC-P, IP, WB

Host Organism

goat

Vendor

Abcam

Paneth Cell Multipotency Induced by Notch Activation following Injury.

  • Yu S
  • Cell Stem Cell
  • 2018 Jul 5

Literature context:


Abstract:

Paneth cells are post-mitotic intestinal epithelial cells supporting the stem cell niche and mucosal immunity. Paneth cell pathologies are observed in various gastrointestinal diseases, but their plasticity and response to genomic and environmental challenges remain unclear. Using a knockin allele engineered at the mouse Lyz1 locus, we performed detailed Paneth cell-lineage tracing. Irradiation induced a subset of Paneth cells to proliferate and differentiate into villus epithelial cells. RNA sequencing (RNA-seq) revealed that Paneth cells sorted from irradiated mice acquired a stem cell-like transcriptome; when cultured in vitro, these individual Paneth cells formed organoids. Irradiation activated Notch signaling, and forced expression of Notch intracellular domain (NICD) in Paneth cells, but not Wnt/β-catenin pathway activation, induced their dedifferentiation. This study documents Paneth cell plasticity, particularly their ability to participate in epithelial replenishment following stem cell loss, adding to a growing body of knowledge detailing the molecular pathways controlling injury-induced regeneration.

Funding information:
  • NCI NIH HHS - R01 CA190558()
  • NCI NIH HHS - R21 CA178599()
  • NCRR NIH HHS - S10 RR027022()
  • NIAMS NIH HHS - R01AR055926(United States)
  • NIDDK NIH HHS - R01 DK102934()
  • NIDDK NIH HHS - R03 DK093809()
  • NIH HHS - S10 OD018103()

Astrocytic signaling supports hippocampal-prefrontal theta synchronization and cognitive function.

  • Sardinha VM
  • Glia
  • 2018 Jun 5

Literature context:


Abstract:

Astrocytes interact with neurons at the cellular level through modulation of synaptic formation, maturation, and function, but the impact of such interaction into behavior remains unclear. Here, we studied the dominant negative SNARE (dnSNARE) mouse model to dissect the role of astrocyte-derived signaling in corticolimbic circuits, with implications for cognitive processing. We found that the blockade of gliotransmitter release in astrocytes triggers a critical desynchronization of neural theta oscillations between dorsal hippocampus and prefrontal cortex. Moreover, we found a strong cognitive impairment in tasks depending on this network. Importantly, the supplementation with d-serine completely restores hippocampal-prefrontal theta synchronization and rescues the spatial memory and long-term memory of dnSNARE mice. We provide here novel evidence of long distance network modulation by astrocytes, with direct implications to cognitive function.

Regulation of Epithelial Plasticity Determines Metastatic Organotropism in Pancreatic Cancer.

  • Reichert M
  • Dev. Cell
  • 2018 Jun 18

Literature context:


Abstract:

The regulation of metastatic organotropism in pancreatic ductal a denocarcinoma (PDAC) remains poorly understood. We demonstrate, using multiple mouse models, that liver and lung metastatic organotropism is dependent upon p120catenin (p120ctn)-mediated epithelial identity. Mono-allelic p120ctn loss accelerates KrasG12D-driven pancreatic cancer formation and liver metastasis. Importantly, one p120ctn allele is sufficient for E-CADHERIN-mediated cell adhesion. By contrast, cells with bi-allelic p120ctn loss demonstrate marked lung organotropism; however, rescue with p120ctn isoform 1A restores liver metastasis. In a p120ctn-independent PDAC model, mosaic loss of E-CADHERIN expression reveals selective pressure for E-CADHERIN-positive liver metastasis and E-CADHERIN-negative lung metastasis. Furthermore, human PDAC and liver metastases support the premise that liver metastases exhibit predominantly epithelial characteristics. RNA-seq demonstrates differential induction of pathways associated with metastasis and epithelial-to-mesenchymal transition in p120ctn-deficient versus p120ctn-wild-type cells. Taken together, P120CTN and E-CADHERIN mediated epithelial plasticity is an addition to the conceptual framework underlying metastatic organotropism in pancreatic cancer.

Funding information:
  • NCI NIH HHS - F30 CA180601()
  • NCI NIH HHS - F32 CA221094()
  • NIDDK NIH HHS - P30 DK050306()
  • NIDDK NIH HHS - R01 DK060694()
  • NIDDK NIH HHS - R21DK090778(United States)

EMT Subtype Influences Epithelial Plasticity and Mode of Cell Migration.

  • Aiello NM
  • Dev. Cell
  • 2018 Jun 18

Literature context:


Abstract:

Epithelial-mesenchymal transition (EMT) is strongly implicated in tumor cell invasion and metastasis. EMT is thought to be regulated primarily at the transcriptional level through the repressive activity of EMT transcription factors. However, these classical mechanisms have been parsed out almost exclusively in vitro, leaving questions about the programs driving EMT in physiological contexts. Here, using a lineage-labeled mouse model of pancreatic ductal adenocarcinoma to study EMT in vivo, we found that most tumors lose their epithelial phenotype through an alternative program involving protein internalization rather than transcriptional repression, resulting in a "partial EMT" phenotype. Carcinoma cells utilizing this program migrate as clusters, contrasting with the single-cell migration pattern associated with traditionally defined EMT mechanisms. Moreover, many breast and colorectal cancer cell lines utilize this alternative program to undergo EMT. Collectively, these results suggest that carcinoma cells have different ways of losing their epithelial program, resulting in distinct modes of invasion and dissemination.

Funding information:
  • Medical Research Council - G0600329(United Kingdom)
  • NCI NIH HHS - F31 CA177163()
  • NCI NIH HHS - K12 CA076931()
  • NCI NIH HHS - P30 CA016087()
  • NCI NIH HHS - R01 CA169123()
  • NIDDK NIH HHS - K08 DK109492()
  • NIDDK NIH HHS - P30 DK050306()

Chrna5-expressing neurons in the interpeduncular nucleus mediate aversion primed by prior stimulation or nicotine exposure.

  • Morton G
  • J. Neurosci.
  • 2018 Jun 28

Literature context:


Abstract:

Genetic studies have shown an association between smoking and variation at the CHRNA5/A3/B4 gene locus, encoding the α5, α3 and β4 nicotinic receptor subunits. The α5 receptor has been specifically implicated because smoking-associated haplotypes contain a coding variant in the CHRNA5 gene. The Chrna5/a3/b4 locus is conserved in rodents, and the restricted expression of these subunits suggests neural pathways through which the reinforcing and aversive properties of nicotine may be mediated. Here we show that in the interpeduncular nucleus (IP), the site of the highest Chrna5 mRNA expression in rodents, electrophysiological responses to nicotinic acetylcholine receptor stimulation are markedly reduced in α5 null mice. In this regard, we find IP neurons differ markedly from their upstream ventral medial habenula cholinergic partners, which appear unaltered by loss of α5. To probe the functional role of α5-containing IP neurons, we used BAC recombineering to generate transgenic mice expressing Cre-recombinase from the Chrna5 locus. Reporter expression driven by Chrna5Cre demonstrates that transcription of Chrna5 is regulated independently from the Chrna3/b4 genes, transcribed on the opposite strand. Chrna5-expressing IP neurons are GABAergic and project to distant targets in the mesopontine raphe and tegmentum, rather than forming local circuits. Optogenetic stimulation of Chrna5-expressing IP neurons failed to elicit physical manifestations of withdrawal. However, following recent prior stimulation or exposure to nicotine, IP stimulation becomes aversive. These results using mice of both sexes support the idea that the risk allele of CHRNA5 may increase the drive to smoke via loss of IP-mediated nicotine aversion.SIGNIFICANCE STATEMENTUnderstanding the receptors and neural pathways underlying the reinforcing and aversive effects of nicotine may suggest new treatments for tobacco addiction. Part of the individual variability in smoking is associated with specific forms of the α5 nicotinic receptor subunit gene. Here we show that deletion of the α5 subunit in mice markedly reduces the cellular response to nicotine and acetylcholine in the interpeduncular nucleus (IP). Stimulation of α5-expressing IP neurons using optogenetics is aversive, but this effect requires priming by recent prior stimulation or exposure to nicotine.. These results support the idea that the smoking-associated variant of the α5 gene may increase the drive to smoke via loss of IP-mediated nicotine aversion.

Funding information:
  • NCI NIH HHS - R01CA90446(United States)
  • NIDA NIH HHS - R01 DA035838(United States)
  • NINDS NIH HHS - N01NS02331(United States)

Nucleus Accumbens Microcircuit Underlying D2-MSN-Driven Increase in Motivation.

  • Soares-Cunha C
  • eNeuro
  • 2018 May 22

Literature context:


Abstract:

The nucleus accumbens (NAc) plays a central role in reinforcement and motivation. Around 95% of the NAc neurons are medium spiny neurons (MSNs), divided into those expressing dopamine receptor D1 (D1R) or dopamine receptor D2 (D2R). Optogenetic activation of D2-MSNs increased motivation, whereas inhibition of these neurons produced the opposite effect. Yet, it is still unclear how activation of D2-MSNs affects other local neurons/interneurons or input terminals and how this contributes for motivation enhancement. To answer this question, in this work we combined optogenetic modulation of D2-MSNs with in loco pharmacological delivery of specific neurotransmitter antagonists in rats. First, we showed that optogenetic activation of D2-MSNs increases motivation in a progressive ratio (PR) task. We demonstrated that this behavioral effect relies on cholinergic-dependent modulation of dopaminergic signalling of ventral tegmental area (VTA) terminals, which requires D1R and D2R signalling in the NAc. D2-MSN optogenetic activation decreased ventral pallidum (VP) activity, reducing the inhibitory tone to VTA, leading to increased dopaminergic activity. Importantly, optogenetic activation of D2-MSN terminals in the VP was sufficient to recapitulate the motivation enhancement. In summary, our data suggests that optogenetic stimulation of NAc D2-MSNs indirectly modulates VTA dopaminergic activity, contributing for increased motivation. Moreover, both types of dopamine receptors signalling in the NAc are required in order to produce the positive behavioral effects.

Funding information:
  • NIBIB NIH HHS - R01 EB008281(United States)

Hippo Signaling Plays an Essential Role in Cell State Transitions during Cardiac Fibroblast Development.

  • Xiao Y
  • Dev. Cell
  • 2018 Apr 23

Literature context:


Abstract:

During development, progenitors progress through transition states. The cardiac epicardium contains progenitors of essential non-cardiomyocytes. The Hippo pathway, a kinase cascade that inhibits the Yap transcriptional co-factor, controls organ size in developing hearts. Here, we investigated Hippo kinases Lats1 and Lats2 in epicardial diversification. Epicardial-specific deletion of Lats1/2 was embryonic lethal, and mutant embryos had defective coronary vasculature remodeling. Single-cell RNA sequencing revealed that Lats1/2 mutant cells failed to activate fibroblast differentiation but remained in an intermediate cell state with both epicardial and fibroblast characteristics. Lats1/2 mutant cells displayed an arrested developmental trajectory with persistence of epicardial markers and expanded expression of Yap targets Dhrs3, an inhibitor of retinoic acid synthesis, and Dpp4, a protease that modulates extracellular matrix (ECM) composition. Genetic and pharmacologic manipulation revealed that Yap inhibits fibroblast differentiation, prolonging a subepicardial-like cell state, and promotes expression of matricellular factors, such as Dpp4, that define ECM characteristics.

Funding information:
  • NIAAA NIH HHS - R01 AA020401(United States)

Reducing Pericyte-Derived Scarring Promotes Recovery after Spinal Cord Injury.

  • Dias DO
  • Cell
  • 2018 Mar 22

Literature context:


Abstract:

CNS injury often severs axons. Scar tissue that forms locally at the lesion site is thought to block axonal regeneration, resulting in permanent functional deficits. We report that inhibiting the generation of progeny by a subclass of pericytes led to decreased fibrosis and extracellular matrix deposition after spinal cord injury in mice. Regeneration of raphespinal and corticospinal tract axons was enhanced and sensorimotor function recovery improved following spinal cord injury in animals with attenuated pericyte-derived scarring. Using optogenetic stimulation, we demonstrate that regenerated corticospinal tract axons integrated into the local spinal cord circuitry below the lesion site. The number of regenerated axons correlated with improved sensorimotor function recovery. In conclusion, attenuation of pericyte-derived fibrosis represents a promising therapeutic approach to facilitate recovery following CNS injury.

Funding information:
  • Intramural NIH HHS - Z01 DE000698-10(United States)

Reconstruction of the Human Colon Epithelium In Vivo.

  • Sugimoto S
  • Cell Stem Cell
  • 2018 Feb 1

Literature context:


Abstract:

Genetic lineage tracing has revealed that Lgr5+ murine colon stem cells (CoSCs) rapidly proliferate at the crypt bottom. However, the spatiotemporal dynamics of human CoSCs in vivo have remained experimentally intractable. Here we established an orthotopic xenograft system for normal human colon organoids, enabling stable reconstruction of the human colon epithelium in vivo. Xenografted organoids were prone to displacement by the remaining murine crypts, and this could be overcome by complete removal of the mouse epithelium. Xenografted organoids formed crypt structures distinctively different from surrounding mouse crypts, reflecting their human origin. Lineage tracing using CRISPR-Cas9 to engineer an LGR5-CreER knockin allele demonstrated self-renewal and multipotency of LGR5+ CoSCs. In contrast to the rapidly cycling properties of mouse Lgr5+ CoSCs, human LGR5+ CoSCs were slow-cycling in vivo. This organoid-based orthotopic xenograft model enables investigation of the functional behaviors of human CoSCs in vivo, with potential therapeutic applications in regenerative medicine.

Funding information:
  • NIGMS NIH HHS - 5T32 GM 7288-37(United States)

Visceral endoderm and the primitive streak interact to build the fetal-placental interface of the mouse gastrula.

  • Rodriguez AM
  • Dev. Biol.
  • 2017 Dec 1

Literature context:


Abstract:

Hypoblast/visceral endoderm assists in amniote nutrition, axial positioning and formation of the gut. Here, we provide evidence, currently limited to humans and non-human primates, that hypoblast is a purveyor of extraembryonic mesoderm in the mouse gastrula. Fate mapping a unique segment of axial extraembryonic visceral endoderm associated with the allantoic component of the primitive streak, and referred to as the "AX", revealed that visceral endoderm supplies the placentae with extraembryonic mesoderm. Exfoliation of the AX was dependent upon contact with the primitive streak, which modulated Hedgehog signaling. Resolution of the AX's epithelial-to-mesenchymal transition (EMT) by Hedgehog shaped the allantois into its characteristic projectile and individualized placental arterial vessels. A unique border cell separated the delaminating AX from the yolk sac blood islands which, situated beyond the limit of the streak, were not formed by an EMT. Over time, the AX became the hindgut lip, which contributed extensively to the posterior interface, including both embryonic and extraembryonic tissues. The AX, in turn, imparted antero-posterior (A-P) polarity on the primitive streak and promoted its elongation and differentiation into definitive endoderm. Results of heterotopic grafting supported mutually interactive functions of the AX and primitive streak, showing that together, they self-organized into a complete version of the fetal-placental interface, forming an elongated structure that exhibited A-P polarity and was composed of the allantois, an AX-derived rod-like axial extension reminiscent of the embryonic notochord, the placental arterial vasculature and visceral endoderm/hindgut.

Hemodynamic Forces Sculpt Developing Heart Valves through a KLF2-WNT9B Paracrine Signaling Axis.

  • Goddard LM
  • Dev. Cell
  • 2017 Nov 6

Literature context:


Abstract:

Hemodynamic forces play an essential epigenetic role in heart valve development, but how they do so is not known. Here, we show that the shear-responsive transcription factor KLF2 is required in endocardial cells to regulate the mesenchymal cell responses that remodel cardiac cushions to mature valves. Endocardial Klf2 deficiency results in defective valve formation associated with loss of Wnt9b expression and reduced canonical WNT signaling in neighboring mesenchymal cells, a phenotype reproduced by endocardial-specific loss of Wnt9b. Studies in zebrafish embryos reveal that wnt9b expression is similarly restricted to the endocardial cells overlying the developing heart valves and is dependent upon both hemodynamic shear forces and klf2a expression. These studies identify KLF2-WNT9B signaling as a conserved molecular mechanism by which fluid forces sensed by endothelial cells direct the complex cellular process of heart valve development and suggest that congenital valve defects may arise due to subtle defects in this mechanotransduction pathway.

Funding information:
  • Cancer Research UK - C7845/A10066(United Kingdom)
  • European Research Council - 682938()
  • NHLBI NIH HHS - R01 HL094326()
  • NHLBI NIH HHS - T32 HL007954()

Mouse Cutaneous Melanoma Induced by Mutant BRaf Arises from Expansion and Dedifferentiation of Mature Pigmented Melanocytes.

  • Köhler C
  • Cell Stem Cell
  • 2017 Nov 2

Literature context:


Abstract:

To identify the cells at the origin of melanoma, we combined single-cell lineage-tracing and transcriptomics approaches with time-lapse imaging. A mouse model that recapitulates key histopathological features of human melanomagenesis was created by inducing a BRafV600E-driven melanomagenic program in tail interfollicular melanocytes. Most targeted mature, melanin-producing melanocytes expanded clonally within the epidermis before losing their differentiated features through transcriptional reprogramming and eventually invading the dermis. Tumors did not form within interscales, which contain both mature and dormant amelanotic melanocytes. The hair follicle bulge, which contains melanocyte stem cells, was also refractory to melanomagenesis. These studies identify varying tumor susceptibilities within the melanocytic lineage, highlighting pigment-producing cells as the melanoma cell of origin, and indicate that regional variation in tumor predisposition is dictated by microenvironmental cues rather than intrinsic differences in cellular origin. Critically, this work provides in vivo evidence that differentiated somatic cells can be reprogrammed into cancer initiating cells.

Adult Conditional Knockout of PGC-1α Leads to Loss of Dopamine Neurons.

  • Jiang H
  • eNeuro
  • 2017 Oct 25

Literature context:


Abstract:

Parkinson's disease (PD) is a chronic progressive neurodegenerative disorder. Recent studies have implicated a role for peroxisome proliferator-activated receptor γ coactivator protein-1α (PGC-1α) in PD and in animal or cellular models of PD. The role of PGC-1α in the function and survival of substantia nigra pars compacta (SNpc) dopamine neurons is not clear. Here we find that there are four different PGC-1α isoforms expressed in SH-SY5Y cells, and these four isoforms are expressed across subregions of mouse brain. Adult conditional PGC-1α knock-out mice show a significant loss of dopaminergic neurons that is accompanied by a reduction of dopamine in the striatum. In human PD postmortem tissue from the SNpc, there is a reduction of PGC-1α isoforms and mitochondria markers. Our findings suggest that all four isoforms of PGC-1α are required for the proper expression of mitochondrial proteins in SNpc DA neurons and that PGC-1α is essential for SNpc DA neuronal survival, possibly through the maintenance of mitochondrial function.

Generation of Mouse and Human Organoid-Forming Intestinal Progenitor Cells by Direct Lineage Reprogramming.

  • Miura S
  • Cell Stem Cell
  • 2017 Oct 5

Literature context:


Abstract:

Intestinal organoids hold great promise as a valuable tool for studying and treating intestinal diseases. The currently available sources of human intestinal organoids, tissue fragments or pluripotent stem cells, involve invasive procedures or complex differentiation protocols, respectively. Here, we show that a set of four transcription factors, Hnf4α, Foxa3, Gata6, and Cdx2, can directly reprogram mouse fibroblasts to acquire the identity of fetal intestine-derived progenitor cells (FIPCs). These induced FIPCs (iFIPCs) form spherical organoids that develop into adult-type budding organoids containing cells with intestinal stem cell properties. The resulting stem cells produce all intestinal epithelial cell lineages and undergo self-renewing cell divisions. After transplantation, the induced spherical and budding organoids can reconstitute colonic and intestinal epithelia, respectively. The same combination of four defined transcription factors can also induce human iFIPCs. This alternative approach for producing intestinal organoids may well facilitate application for disease analysis and therapy development.

Inter-dependent apical microtubule and actin dynamics orchestrate centrosome retention and neuronal delamination.

  • Kasioulis I
  • Elife
  • 2017 Oct 23

Literature context:


Abstract:

Detachment of newborn neurons from the neuroepithelium is required for correct neuronal architecture and functional circuitry. This process, also known as delamination, involves adherens-junction disassembly and acto-myosin-mediated abscission, during which the centrosome is retained while apical/ciliary membranes are shed. Cell-biological mechanisms mediating delamination are, however, poorly understood. Using live-tissue and super-resolution imaging, we uncover a centrosome-nucleated wheel-like microtubule configuration, aligned with the apical actin cable and adherens-junctions within chick and mouse neuroepithelial cells. These microtubules maintain adherens-junctions while actin maintains microtubules, adherens-junctions and apical end-foot dimensions. During neuronal delamination, acto-myosin constriction generates a tunnel-like actin-microtubule configuration through which the centrosome translocates. This movement requires inter-dependent actin and microtubule activity, and we identify drebrin as a potential coordinator of these cytoskeletal dynamics. Furthermore, centrosome compromise revealed that this organelle is required for delamination. These findings identify new cytoskeletal configurations and regulatory relationships that orchestrate neuronal delamination and may inform mechanisms underlying pathological epithelial cell detachment.

Heterophilic Type II Cadherins Are Required for High-Magnitude Synaptic Potentiation in the Hippocampus.

  • Basu R
  • Neuron
  • 2017 Sep 27

Literature context:


Abstract:

Hippocampal CA3 neurons form synapses with CA1 neurons in two layers, stratum oriens (SO) and stratum radiatum (SR). Each layer develops unique synaptic properties but molecular mechanisms that mediate these differences are unknown. Here, we show that SO synapses normally have significantly more mushroom spines and higher-magnitude long-term potentiation (LTP) than SR synapses. Further, we discovered that these differences require the Type II classic cadherins, cadherins-6, -9, and -10. Though cadherins typically function via trans-cellular homophilic interactions, our results suggest presynaptic cadherin-9 binds postsynaptic cadherins-6 and -10 to regulate mushroom spine density and high-magnitude LTP in the SO layer. Loss of these cadherins has no effect on the lower-magnitude LTP typically observed in the SR layer, demonstrating that cadherins-6, -9, and -10 are gatekeepers for high-magnitude LTP. Thus, Type II cadherins may uniquely contribute to the specificity and strength of synaptic changes associated with learning and memory.

Funding information:
  • NEI NIH HHS - R01 EY022073()

Sonic Hedgehog switches on Wnt/planar cell polarity signaling in commissural axon growth cones by reducing levels of Shisa2.

  • Onishi K
  • Elife
  • 2017 Sep 8

Literature context:


Abstract:

Commissural axons switch on responsiveness to Wnt attraction during midline crossing and turn anteriorly only after exiting the floor plate. We report here that Sonic Hedgehog (Shh)-Smoothened signaling downregulates Shisa2, which inhibits the glycosylation and cell surface presentation of Frizzled3 in rodent commissural axon growth cones. Constitutive Shisa2 expression causes randomized turning of post-crossing commissural axons along the anterior-posterior (A-P) axis. Loss of Shisa2 led to precocious anterior turning of commissural axons before or during midline crossing. Post-crossing commissural axon turning is completely randomized along the A-P axis when Wntless, which is essential for Wnt secretion, is conditionally knocked out in the floor plate. This regulatory link between Shh and planar cell polarity (PCP) signaling may also occur in other developmental processes.

Collaboration between Distinct Rab Small GTPase Trafficking Circuits Mediates Bacterial Clearance from the Bladder Epithelium.

  • Miao Y
  • Cell Host Microbe
  • 2017 Sep 13

Literature context:


Abstract:

Rab small GTPases control membrane trafficking through effectors that recruit downstream mediators such as motor proteins. Subcellular trafficking typically involves multiple Rabs, with each specific step mediated by a distinct Rab protein. We describe a collaboration between two distinct Rab-protein-orchestrated trafficking circuits in bladder epithelial cells (BECs) that expels intracellular uropathogenic Escherichia coli (UPEC) from their intracellular niche. RAB11a and RAB27b and their trafficking circuitry are simultaneously involved in UPEC expulsion. While RAB11a recruits its effector RAB11FIP3 and cytoskeletal motor Dynein, RAB27b mobilizes the effector MyRIP and motor Myosin VIIa to mediate bacterial expulsion. This collaboration is coordinated by deposition of the exocyst complex on bacteria-containing vesicles, an event triggered by the innate receptor Toll-like receptor 4. Both RAB11a and RAB27b are recruited and activated by the exocyst complex components SEC6/SEC15. Thus, the cell autonomous defense system can mobilize and coalesce multiple subcellular trafficking circuitries to combat infections.

Distinct Mesenchymal Lineages and Niches Promote Epithelial Self-Renewal and Myofibrogenesis in the Lung.

  • Zepp JA
  • Cell
  • 2017 Sep 7

Literature context:


Abstract:

The lung is an architecturally complex organ comprising a heterogeneous mixture of various epithelial and mesenchymal lineages. We use single-cell RNA sequencing and signaling lineage reporters to generate a spatial and transcriptional map of the lung mesenchyme. We find that each mesenchymal lineage has a distinct spatial address and transcriptional profile leading to unique niche regulatory functions. The mesenchymal alveolar niche cell is Wnt responsive, expresses Pdgfrα, and is critical for alveolar epithelial cell growth and self-renewal. In contrast, the Axin2+ myofibrogenic progenitor cell preferentially generates pathologically deleterious myofibroblasts after injury. Analysis of the secretome and receptome of the alveolar niche reveals functional pathways that mediate growth and self-renewal of alveolar type 2 progenitor cells, including IL-6/Stat3, Bmp, and Fgf signaling. These studies define the cellular and molecular framework of lung mesenchymal niches and reveal the functional importance of developmental pathways in promoting self-renewal versus a pathological response to tissue injury.

Intra-lineage Fate Decisions Involve Activation of Notch Receptors Basal to the Midbody in Drosophila Sensory Organ Precursor Cells.

  • Trylinski M
  • Curr. Biol.
  • 2017 Aug 7

Literature context:


Abstract:

Notch receptors regulate cell fate decisions during embryogenesis and throughout adult life. In many cell lineages, binary fate decisions are mediated by directional Notch signaling between the two sister cells produced by cell division. How Notch signaling is restricted to sister cells after division to regulate intra-lineage decision is poorly understood. More generally, where ligand-dependent activation of Notch occurs at the cell surface is not known, as methods to detect receptor activation in vivo are lacking. In Drosophila pupae, Notch signals during cytokinesis to regulate the intra-lineage pIIa/pIIb decision in the sensory organ lineage. Here, we identify two pools of Notch along the pIIa-pIIb interface, apical and basal to the midbody. Analysis of the dynamics of Notch, Delta, and Neuralized distribution in living pupae suggests that ligand endocytosis and receptor activation occur basal to the midbody. Using selective photo-bleaching of GFP-tagged Notch and photo-tracking of photo-convertible Notch, we show that nuclear Notch is indeed produced by receptors located basal to the midbody. Thus, only a specific subset of receptors, located basal to the midbody, contributes to signaling in pIIa. This is the first in vivo characterization of the pool of Notch contributing to signaling. We propose a simple mechanism of cell fate decision based on intra-lineage signaling: ligands and receptors localize during cytokinesis to the new cell-cell interface, thereby ensuring signaling between sister cells, hence intra-lineage fate decision.

Exosome RNA Unshielding Couples Stromal Activation to Pattern Recognition Receptor Signaling in Cancer.

  • Nabet BY
  • Cell
  • 2017 Jul 13

Literature context:


Abstract:

Interactions between stromal fibroblasts and cancer cells generate signals for cancer progression, therapy resistance, and inflammatory responses. Although endogenous RNAs acting as damage-associated molecular patterns (DAMPs) for pattern recognition receptors (PRRs) may represent one such signal, these RNAs must remain unrecognized under non-pathological conditions. We show that triggering of stromal NOTCH-MYC by breast cancer cells results in a POL3-driven increase in RN7SL1, an endogenous RNA normally shielded by RNA binding proteins SRP9/14. This increase in RN7SL1 alters its stoichiometry with SRP9/14 and generates unshielded RN7SL1 in stromal exosomes. After exosome transfer to immune cells, unshielded RN7SL1 drives an inflammatory response. Upon transfer to breast cancer cells, unshielded RN7SL1 activates the PRR RIG-I to enhance tumor growth, metastasis, and therapy resistance. Corroborated by evidence from patient tumors and blood, these results demonstrate that regulation of RNA unshielding couples stromal activation with deployment of RNA DAMPs that promote aggressive features of cancer. VIDEO ABSTRACT.

Mex3a Marks a Slowly Dividing Subpopulation of Lgr5+ Intestinal Stem Cells.

  • Barriga FM
  • Cell Stem Cell
  • 2017 Jun 1

Literature context:


Abstract:

Highly proliferative Lgr5+ stem cells maintain the intestinal epithelium and are thought to be largely homogeneous. Although quiescent intestinal stem cell (ISC) populations have been described, the identity and features of such a population remain controversial. Here we report unanticipated heterogeneity within the Lgr5+ ISC pool. We found that expression of the RNA-binding protein Mex3a labels a slowly cycling subpopulation of Lgr5+ ISCs that contribute to all intestinal lineages with distinct kinetics. Single-cell transcriptome profiling revealed that Lgr5+ cells adopt two discrete states, one of which is defined by a Mex3a expression program and relatively low levels of proliferation genes. During homeostasis, Mex3a+ cells continually shift into the rapidly dividing, self-renewing ISC pool. Chemotherapy and radiation preferentially target rapidly dividing Lgr5+ cells but spare the Mex3a-high/Lgr5+ population, helping to promote regeneration of the intestinal epithelium following toxic insults. Thus, Mex3a defines a reserve-like ISC population within the Lgr5+ compartment.

Hoxa9 and Meis1 Cooperatively Induce Addiction to Syk Signaling by Suppressing miR-146a in Acute Myeloid Leukemia.

  • Mohr S
  • Cancer Cell
  • 2017 Apr 10

Literature context:


Abstract:

The transcription factor Meis1 drives myeloid leukemogenesis in the context of Hox gene overexpression but is currently considered undruggable. We therefore investigated whether myeloid progenitor cells transformed by Hoxa9 and Meis1 become addicted to targetable signaling pathways. A comprehensive (phospho)proteomic analysis revealed that Meis1 increased Syk protein expression and activity. Syk upregulation occurs through a Meis1-dependent feedback loop. By dissecting this loop, we show that Syk is a direct target of miR-146a, whose expression is indirectly regulated by Meis1 through the transcription factor PU.1. In the context of Hoxa9 overexpression, Syk signaling induces Meis1, recapitulating several leukemogenic features of Hoxa9/Meis1-driven leukemia. Finally, Syk inhibition disrupts the identified regulatory loop, prolonging survival of mice with Hoxa9/Meis1-driven leukemia.

Funding information:
  • NCI NIH HHS - R01 CA140292()
  • NCI NIH HHS - R35 CA210030()

Serum Proteases Potentiate BMP-Induced Cell Cycle Re-entry of Dedifferentiating Muscle Cells during Newt Limb Regeneration.

  • Wagner I
  • Dev. Cell
  • 2017 Mar 27

Literature context:


Abstract:

Limb amputation in the newt induces myofibers to dedifferentiate and re-enter the cell cycle to generate proliferative myogenic precursors in the regeneration blastema. Here we show that bone morphogenetic proteins (BMPs) and mature BMPs that have been further cleaved by serum proteases induce cell cycle entry by dedifferentiating newt muscle cells. Protease-activated BMP4/7 heterodimers that are present in serum strongly induced myotube cell cycle re-entry with protease cleavage yielding a 30-fold potency increase of BMP4/7 compared with canonical BMP4/7. Inhibition of BMP signaling via muscle-specific dominant-negative receptor expression reduced cell cycle entry in vitro and in vivo. In vivo inhibition of serine protease activity depressed cell cycle re-entry, which in turn was rescued by cleaved-mimic BMP. This work identifies a mechanism of BMP activation that generates blastema cells from differentiated muscle.

Distinct Hippocampal Pathways Mediate Dissociable Roles of Context in Memory Retrieval.

  • Xu C
  • Cell
  • 2016 Nov 3

Literature context:


Abstract:

Memories about sensory experiences are tightly linked to the context in which they were formed. Memory contextualization is fundamental for the selection of appropriate behavioral reactions needed for survival, yet the underlying neuronal circuits are poorly understood. By combining trans-synaptic viral tracing and optogenetic manipulation, we found that the ventral hippocampus (vHC) and the amygdala, two key brain structures encoding context and emotional experiences, interact via multiple parallel pathways. A projection from the vHC to the basal amygdala mediates fear behavior elicited by a conditioned context, whereas a parallel projection from a distinct subset of vHC neurons onto midbrain-projecting neurons in the central amygdala is necessary for context-dependent retrieval of cued fear memories. Our findings demonstrate that two fundamentally distinct roles of context in fear memory retrieval are processed by distinct vHC output pathways, thereby allowing for the formation of robust contextual fear memories while preserving context-dependent behavioral flexibility.

Tau and spectraplakins promote synapse formation and maintenance through Jun kinase and neuronal trafficking.

  • Voelzmann A
  • Elife
  • 2016 Aug 8

Literature context:


Abstract:

The mechanisms regulating synapse numbers during development and ageing are essential for normal brain function and closely linked to brain disorders including dementias. Using Drosophila, we demonstrate roles of the microtubule-associated protein Tau in regulating synapse numbers, thus unravelling an important cellular requirement of normal Tau. In this context, we find that Tau displays a strong functional overlap with microtubule-binding spectraplakins, establishing new links between two different neurodegenerative factors. Tau and the spectraplakin Short Stop act upstream of a three-step regulatory cascade ensuring adequate delivery of synaptic proteins. This cascade involves microtubule stability as the initial trigger, JNK signalling as the central mediator, and kinesin-3 mediated axonal transport as the key effector. This cascade acts during development (synapse formation) and ageing (synapse maintenance) alike. Therefore, our findings suggest novel explanations for intellectual disability in Tau deficient individuals, as well as early synapse loss in dementias including Alzheimer's disease.

Expression Analysis of CB2-GFP BAC Transgenic Mice.

  • Schmöle AC
  • PLoS ONE
  • 2015 Sep 26

Literature context:


Abstract:

The endocannabinoid system (ECS) is a retrograde messenger system, consisting of lipid signaling molecules that bind to at least two G-protein-coupled receptors, Cannabinoid receptor 1 and 2 (CB1 and 2). As CB2 is primarily expressed on immune cells such as B cells, T cells, macrophages, dendritic cells, and microglia, it is of great interest how CB2 contributes to immune cell development and function in health and disease. Here, understanding the mechanisms of CB2 involvement in immune-cell function as well as the trafficking and regulation of CB2 expressing cells are crucial issues. Up to now, CB2 antibodies produce unclear results, especially those targeting the murine protein. Therefore, we have generated BAC transgenic GFP reporter mice (CB2-GFPTg) to trace CB2 expression in vitro and in situ. Those mice express GFP under the CB2 promoter and display GFP expression paralleling CB2 expression on the transcript level in spleen, thymus and brain tissue. Furthermore, by using fluorescence techniques we show that the major sources for GFP-CB2 expression are B cells in spleen and blood and microglia in the brain. This novel CB2-GFP transgenic reporter mouse line represents a powerful resource to study CB2 expression in different cell types. Furthermore, it could be used for analyzing CB2-mediated mobilization and trafficking of immune cells as well as studying the fate of recruited immune cells in models of acute and chronic inflammation.

Funding information:
  • NCRR NIH HHS - P40-RR17072(United States)
  • NIAAA NIH HHS - R01 AA017413(United States)

Duodenal CCK cells from male mice express multiple hormones including ghrelin.

  • Sykaras AG
  • Endocrinology
  • 2014 Sep 25

Literature context:


Abstract:

Enteroendocrine (EEC) cells have a pivotal role in intestinal nutrient sensing and release hormones that orchestrate food digestion and regulate appetite. EEC cells are found scattered throughout the intestine and have typically been classified based on the primary hormone they contain. I cells represent a subset of EEC cells that secrete cholecystokinin (CCK) and are mainly localized to the duodenum. Recent studies have shown that I cells express mRNAs encoding several gut hormones. In this study, we investigated the hormonal profile of murine fluorescence-activated cell sorting-sorted duodenal I cells using semiquantitative RT-PCR, liquid chromatography tandem mass spectrometry, and immunostaining methods. We report that I cells are enriched in mRNA transcripts encoding CCK and also other key gut hormones, including neurotensin, glucose-dependent insulinotropic peptide (GIP), secretin, peptide YY, proglucagon, and ghrelin (Ghrl). Furthermore, liquid chromatography tandem mass spectrometry analysis of fluorescence-activated cell sorting-purified I cells and immunostaining confirmed the presence of these gut hormones in duodenal I cells. Immunostaining highlighted that subsets of I cells in both crypts and villi coexpress differential amounts of CCK, Ghrl, GIP, or peptide YY, indicating that a proportion of I cells contain several hormones during maturation and when fully differentiated. Our results reveal that although I cells express several key gut hormones, including GIP or proglucagon, and thus have a considerable overlap with classically defined K and L cells, approximately half express Ghrl, suggesting a potentially important subset of duodenal EEC cells that require further consideration.

Funding information:
  • Howard Hughes Medical Institute - R01NS036715(United States)

Establishment of mice expressing EGFP in the placode-derived inner ear sensory cell lineage and FACS-array analysis focused on the regional specificity of the otocyst.

  • Fujimoto C
  • J. Comp. Neurol.
  • 2010 Dec 1

Literature context:


Abstract:

In this study, we established a novel enhanced green fluorescent protein (EGFP) reporter mouse line that enables the visualization of the placode-derived inner ear sensory cell lineage. EGFP was initially expressed in the otic placode and throughout its differentiation process into the inner ear sensory patches. At embryonic day 10.5 (E10.5), EGFP was expressed in the ventral and dorsomedial region of the otocyst. These regions could mainly give rise to the cochlea, including the organ of Corti, and the saccule, including the macula and the endolymphatic duct. The region could also give rise to cells that will develop as either prosensory cells or statoacoustic ganglion neuroblasts. By using this line and fluorescence-activated cell sorting (FACS)-array technology, we developed a new gene expression profile of the regional specificity of the otocyst. EGFP-positive regions include the Otx1-positive region, which could be clearly distinguished from EGFP-negative regions. The signal log ratio of microarray data showed high efficiency in predicting the genes expressed mainly in the ventral and/or dorsomedial otocyst and the data could be mined to uncover many novel genes involved in inner ear morphogenesis and cell fate regulation. Additionally, these data suggest that some novel genes enriched in EGFP-positive regions may be potentially involved in human congenital sensorineural hearing loss. This reporter line could play important roles in the use of animal models for detailed analysis of the differentiation process into the sensory patches and the identification of regional-specific gene networks and novel gene functions in the developing inner ear.

Funding information:
  • NHLBI NIH HHS - U54HL108460(United States)
  • NIMH NIH HHS - R01-MH046516(United States)