Neuronal Activity Promotes Glioma Growth through Neuroligin-3 Secretion

Active neurons exert a mitogenic effect on normal neural precursor and oligodendroglial precursor cells, the putative cellular origins of high-grade glioma (HGG). By using optogenetic control of cortical neuronal activity in a patient-derived pediatric glioblastoma xenograft model, we demonstrate that active neurons similarly promote HGG proliferation and growth in vivo. Conditioned medium from optogenetically stimulated cortical slices promoted proliferation of pediatric and adult patient-derived HGG cultures, indicating secretion of activity-regulated mitogen(s). The synaptic protein neuroligin-3 (NLGN3) was identified as the leading candidate mitogen, and soluble NLGN3 was sufficient and necessary to promote robust HGG cell proliferation. NLGN3 induced PI3K-mTOR pathway activity and feedforward expression of NLGN3 in glioma cells. NLGN3 expression levels in human HGG negatively correlated with patient overall survival. These findings indicate the important role of active neurons in the brain tumor microenvironment and identify secreted NLGN3 as an unexpected mechanism promoting neuronal activity-regulated cancer growth.

[1]  Tom R. Gaunt,et al.  Predicting the Functional, Molecular, and Phenotypic Consequences of Amino Acid Substitutions using Hidden Markov Models , 2012, Human mutation.

[2]  D. Steindler,et al.  Chondroitin Sulfate Proteoglycans Potently Inhibit Invasion and Serve as a Central Organizer of the Brain Tumor Microenvironment , 2013, The Journal of Neuroscience.

[3]  Thomas C. Südhof,et al.  Neuroligins Determine Synapse Maturation and Function , 2006, Neuron.

[4]  T. Südhof,et al.  Neuroligin-2 Deletion Selectively Decreases Inhibitory Synaptic Transmission Originating from Fast-Spiking but Not from Somatostatin-Positive Interneurons , 2009, The Journal of Neuroscience.

[5]  T. Maniatis,et al.  An RNA-Sequencing Transcriptome and Splicing Database of Glia, Neurons, and Vascular Cells of the Cerebral Cortex , 2014, The Journal of Neuroscience.

[6]  T. Mikkelsen,et al.  GRP78 is overexpressed in glioblastomas and regulates glioma cell growth and apoptosis. , 2008, Neuro-oncology.

[7]  Noemi Andor,et al.  Asymmetry-defective oligodendrocyte progenitors are glioma precursors. , 2011, Cancer cell.

[8]  Karl Deisseroth,et al.  Optogenetics in Neural Systems , 2011, Neuron.

[9]  Robertson Craig,et al.  Open source system for analyzing, validating, and storing protein identification data. , 2004, Journal of proteome research.

[10]  William Stafford Noble,et al.  Estimating relative abundances of proteins from shotgun proteomics data , 2012, BMC Bioinformatics.

[11]  David T. W. Jones,et al.  Driver mutations in histone H3.3 and chromatin remodelling genes in paediatric glioblastoma , 2012, Nature.

[12]  Christina S. Leslie,et al.  CSF-1R inhibition alters macrophage polarization and blocks glioma progression , 2013, Nature Medicine.

[13]  Johan Pallud,et al.  NG2+/Olig2+ Cells are the Major Cycle‐Related Cell Population of the Adult Human Normal Brain , 2010, Brain pathology.

[14]  M. Berger,et al.  Non-stem cell origin for oligodendroglioma. , 2010, Cancer cell.

[15]  I. Weissman,et al.  Hedgehog-responsive candidate cell of origin for diffuse intrinsic pontine glioma , 2011, Proceedings of the National Academy of Sciences.

[16]  G. T. Bowden,et al.  The role of PI 3-kinase in the UVB-induced expression of c-fos , 2002, Oncogene.

[17]  David T. W. Jones,et al.  K27M mutation in histone H3.3 defines clinically and biologically distinct subgroups of pediatric diffuse intrinsic pontine gliomas , 2012, Acta Neuropathologica.

[18]  Mihaela E. Sardiu,et al.  Enriching quantitative proteomics with SIN , 2010, Nature Biotechnology.

[19]  David T. W. Jones,et al.  Hotspot mutations in H3F3A and IDH1 define distinct epigenetic and biological subgroups of glioblastoma. , 2012, Cancer cell.

[20]  Harald Sontheimer,et al.  Glutamate Release by Primary Brain Tumors Induces Epileptic Activity , 2011, Nature Medicine.

[21]  Keith L Ligon,et al.  The central nervous system-restricted transcription factor Olig2 opposes p53 responses to genotoxic damage in neural progenitors and malignant glioma. , 2011, Cancer cell.

[22]  P. Somogyi,et al.  Glutamatergic synapses on oligodendrocyte precursor cells in the hippocampus , 2000, Nature.

[23]  P. Mallick,et al.  Investigation of acquired resistance to EGFR-targeted therapies in lung cancer using cDNA microarrays. , 2012, Methods in molecular biology.

[24]  Alexey I Nesvizhskii,et al.  Empirical statistical model to estimate the accuracy of peptide identifications made by MS/MS and database search. , 2002, Analytical chemistry.

[25]  Y. Yoshida,et al.  Blockage of Ca(2+)-permeable AMPA receptors suppresses migration and induces apoptosis in human glioblastoma cells. , 2002, Nature medicine.

[26]  H. Tomita,et al.  Denervation suppresses gastric tumorigenesis , 2014, Science Translational Medicine.

[27]  Dong-bao Chen,et al.  Epidermal growth factor induces c-fos and c-jun mRNA via Raf-1/MEK1/ERK-dependent and -independent pathways in bovine luteal cells , 2003, Molecular and Cellular Endocrinology.

[28]  W. C. Hall,et al.  High-speed mapping of synaptic connectivity using photostimulation in Channelrhodopsin-2 transgenic mice , 2007, Proceedings of the National Academy of Sciences.

[29]  F. Kirchhoff,et al.  GABAA‐receptor expression in glioma cells is triggered by contact with neuronal cells , 2001, The European journal of neuroscience.

[30]  Benjamin R. Arenkiel,et al.  In Vivo Light-Induced Activation of Neural Circuitry in Transgenic Mice Expressing Channelrhodopsin-2 , 2007, Neuron.

[31]  Michael E. Greenberg,et al.  Stimulation of 3T3 cells induces transcription of the c-fos proto-oncogene , 1984, Nature.

[32]  Christopher W Mount,et al.  Neuronal Activity Promotes Oligodendrogenesis and Adaptive Myelination in the Mammalian Brain , 2014, Science.

[33]  Christos G. Gkogkas,et al.  Autism-related deficits via dysregulated eIF4E-dependent translational control , 2012, Nature.

[34]  Michael K. Coleman,et al.  Analyzing chromatin remodeling complexes using shotgun proteomics and normalized spectral abundance factors. , 2006, Methods.

[35]  D. Bergles,et al.  Neuron-glia synapses in the brain , 2010, Brain Research Reviews.

[36]  R. DePinho,et al.  Neuroplastic changes occur early in the development of pancreatic ductal adenocarcinoma. , 2014, Cancer research.

[37]  J. Yates,et al.  A model for random sampling and estimation of relative protein abundance in shotgun proteomics. , 2004, Analytical chemistry.

[38]  M. Hediger,et al.  Knockout of Glutamate Transporters Reveals a Major Role for Astroglial Transport in Excitotoxicity and Clearance of Glutamate , 1996, Neuron.

[39]  R. Verhaak,et al.  Transformation of quiescent adult oligodendrocyte precursor cells into malignant glioma through a multistep reactivation process , 2014, Proceedings of the National Academy of Sciences.

[40]  Yuan Qi,et al.  Integrated Genomic Analysis Identifies Clinically Relevant Subtypes of Glioblastoma Characterized by Abnormalities in PDGFRA , IDH 1 , EGFR , and NF 1 Citation Verhaak , 2010 .

[41]  T. Südhof.  Neuroligins and neurexins link synaptic function to cognitive disease , 2008, Nature.

[42]  Mark D Robinson,et al.  edgeR for differential RNA-seq and ChIP-seq analysis: an application to stem cell biology. , 2014, Methods in molecular biology.

[43]  R. Aebersold,et al.  A statistical model for identifying proteins by tandem mass spectrometry. , 2003, Analytical chemistry.

[44]  Benjamin E. Gross,et al.  Integrative Analysis of Complex Cancer Genomics and Clinical Profiles Using the cBioPortal , 2013, Science Signaling.

[45]  Ruedi Aebersold,et al.  Statistical protein quantification and significance analysis in label-free LC-MS experiments with complex designs , 2012, BMC Bioinformatics.

[46]  H. Scherer.  Structural Development in Gliomas , 1938 .

[47]  Li Ding,et al.  Somatic Histone H3 Alterations in Paediatric Diffuse Intrinsic Pontine Gliomas and Non-Brainstem Glioblastomas , 2012, Nature Genetics.

[48]  K. Deisseroth,et al.  Millisecond-timescale, genetically targeted optical control of neural activity , 2005, Nature Neuroscience.

[49]  George Paxinos,et al.  The Mouse Brain in Stereotaxic Coordinates , 2001 .

[50]  S. Gabriel,et al.  Integrated genomic analysis identifies clinically relevant subtypes of glioblastoma characterized by abnormalities in PDGFRA, IDH1, EGFR, and NF1. , 2010, Cancer cell.

[51]  Veronika A. Glukhova,et al.  Contribution of protein fractionation to depth of analysis of the serum and plasma proteomes. , 2007, Journal of proteome research.

[52]  Mark D. Robinson,et al.  edgeR: a Bioconductor package for differential expression analysis of digital gene expression data , 2009, Bioinform..

[53]  R. Aebersold,et al.  A High-Confidence Human Plasma Proteome Reference Set with Estimated Concentrations in PeptideAtlas* , 2011, Molecular & Cellular Proteomics.

[54]  H. Sontheimer,et al.  Bradykinin enhances invasion of malignant glioma into the brain parenchyma by inducing cells to undergo amoeboid migration , 2014, The Journal of physiology.

[55]  Adam Rauch,et al.  Computational Proteomics Analysis System (CPAS): an extensible, open-source analytic system for evaluating and publishing proteomic data and high throughput biological experiments. , 2006, Journal of proteome research.

[56]  H. Kettenmann,et al.  The brain tumor microenvironment , 2011, Glia.

[57]  Y. Yoshida,et al.  Ca2+-Permeable AMPA Receptors Regulate Growth of Human Glioblastoma via Akt Activation , 2007, The Journal of Neuroscience.

[58]  T. Südhof,et al.  Structures, Alternative Splicing, and Neurexin Binding of Multiple Neuroligins (*) , 1996, The Journal of Biological Chemistry.

[59]  Shawn M. Gillespie,et al.  Single-cell RNA-seq highlights intratumoral heterogeneity in primary glioblastoma , 2014, Science.

[60]  M. Wegner,et al.  Mutual Antagonism Between Sox10 and NFIA Regulates Diversification of Glial Lineages and Glioma Sub-Types , 2014, Nature Neuroscience.

[61]  Y. Wang,et al.  Expression of mutant p53 proteins implicates a lineage relationship between neural stem cells and malignant astrocytic glioma in a murine model. , 2009, Cancer cell.

[62]  Frank McCormick,et al.  EGFR Signals to mTOR Through PKC and Independently of Akt in Glioma , 2009, Science Signaling.

[63]  Thomas Bourgeron,et al.  Mutations of the X-linked genes encoding neuroligins NLGN3 and NLGN4 are associated with autism , 2003, Nature Genetics.

[64]  L. Luo,et al.  Mosaic Analysis with Double Markers Reveals Tumor Cell of Origin in Glioma , 2011, Cell.

[65]  T. Südhof,et al.  Cartography of neurexins: More than 1000 isoforms generated by alternative splicing and expressed in distinct subsets of neurons , 1995, Neuron.

[66]  V. Berezin,et al.  Neuroligin‐1 induces neurite outgrowth through interaction with neurexin‐1β and activation of fibroblast growth factor receptor‐1 , 2012, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[67]  S. Torp,et al.  The clinical value of Ki-67/MIB-1 labeling index in human astrocytomas , 2008, Pathology & Oncology Research.

[68]  Jacqueline Blundell,et al.  A Neuroligin-3 Mutation Implicated in Autism Increases Inhibitory Synaptic Transmission in Mice , 2007, Science.

[69]  H. Kettenmann,et al.  Functional GABAA receptors on human glioma cells , 1998, The European journal of neuroscience.

[70]  Harald Sontheimer,et al.  A neurocentric perspective on glioma invasion , 2014, Nature Reviews Neuroscience.

[71]  Patrick E. Rothwell,et al.  Autism-Associated Neuroligin-3 Mutations Commonly Impair Striatal Circuits to Boost Repetitive Behaviors , 2014, Cell.

[72]  William Stafford Noble,et al.  Crux: Rapid Open Source Protein Tandem Mass Spectrometry Analysis , 2014, Journal of proteome research.

[73]  E. Marcotte,et al.  Calculating absolute and relative protein abundance from mass spectrometry-based protein expression data , 2008, Nature Protocols.

[74]  Adrian V. Lee,et al.  Proteomic and transcriptomic profiling reveals a link between the PI3K pathway and lower estrogen-receptor (ER) levels and activity in ER+ breast cancer , 2010, Breast Cancer Research.

[75]  Philip Gerlee,et al.  The model muddle: in search of tumor growth laws. , 2012, Cancer research.

[76]  N. Matsuki,et al.  Activity-Dependent Proteolytic Cleavage of Neuroligin-1 , 2012, Neuron.

[77]  T. Lagerweij,et al.  Human pontine glioma cells can induce murine tumors , 2014, Acta Neuropathologica.

[78]  M. Ehlers,et al.  Transsynaptic Signaling by Activity-Dependent Cleavage of Neuroligin-1 , 2012, Neuron.

[79]  Harald Sontheimer,et al.  Human glioma cells induce hyperexcitability in cortical networks , 2012, Epilepsia.

[80]  Jonathan Katz,et al.  Installation and Use of LabKey Server for Proteomics , 2011, Current protocols in bioinformatics.

[81]  S. Freedland,et al.  Autonomic Nerve Development Contributes to Prostate Cancer Progression , 2013, Science.

[82]  F. Bussolino,et al.  Neuroligin 1 Induces Blood Vessel Maturation by Cooperating with the α6 Integrin* , 2014, The Journal of Biological Chemistry.