Deletion of Transglutaminase 2 from Mouse Astrocytes Significantly Improves Their Ability to Promote Neurite Outgrowth on an Inhibitory Matrix

Astrocytes are the primary support cells of the central nervous system (CNS) that help maintain the energetic requirements and homeostatic environment of neurons. CNS injury causes astrocytes to take on reactive phenotypes with an altered overall function that can range from supportive to harmful for recovering neurons. The characterization of reactive astrocyte populations is a rapidly developing field, and the underlying factors and signaling pathways governing which type of reactive phenotype that astrocytes take on are poorly understood. Our previous studies suggest that transglutaminase 2 (TG2) has an important role in determining the astrocytic response to injury. Selectively deleting TG2 from astrocytes improves functional outcomes after CNS injury and causes widespread changes in gene regulation, which is associated with its nuclear localization. To begin to understand how TG2 impacts astrocytic function, we used a neuron-astrocyte co-culture paradigm to compare the effects of TG2−/− and wild-type (WT) mouse astrocytes on neurite outgrowth and synapse formation. Neurons were grown on a control substrate or an injury-simulating matrix comprised of inhibitory chondroitin sulfate proteoglycans (CSPGs). Compared to WT astrocytes, TG2−/− astrocytes supported neurite outgrowth to a significantly greater extent only on the CSPG matrix, while synapse formation assays showed mixed results depending on the pre- and post-synaptic markers analyzed. We hypothesize that TG2 regulates the supportive functions of astrocytes in injury conditions by modulating gene expression through interactions with transcription factors and transcription complexes. Based on the results of a previous yeast two-hybrid screen for TG2 interactors, we further investigated the interaction of TG2 with Zbtb7a, a ubiquitously expressed transcription factor. Co-immunoprecipitation and colocalization analyses confirmed the interaction of TG2 and Zbtb7a in the nucleus of astrocytes. Overexpression or knockdown of Zbtb7a levels in WT and TG2−/− astrocytes revealed that Zbtb7a robustly influenced astrocytic morphology and the ability of astrocytes to support neuronal outgrowth, which was significantly modulated by the presence of TG2. These findings support our hypothesis that astrocytic TG2 acts as a transcriptional regulator to influence astrocytic function, with greater influence under injury conditions that increase its expression, and Zbtb7a likely contributes to the overall effects observed with astrocytic TG2 deletion.

[1]  Y. Ao,et al.  Divergent transcriptional regulation of astrocyte reactivity across disorders , 2022, Nature.

[2]  B. Barres,et al.  Neurotoxic reactive astrocytes induce cell death via saturated lipids , 2021, Nature.

[3]  G. Johnson,et al.  Deletion or Inhibition of Astrocytic Transglutaminase 2 Promotes Functional Recovery after Spinal Cord Injury , 2021, bioRxiv.

[4]  K. Sakamoto,et al.  Axonal Regeneration by Glycosaminoglycan , 2021, Frontiers in Cell and Developmental Biology.

[5]  Kira E. Poskanzer,et al.  Reactive astrocyte nomenclature, definitions, and future directions , 2021, Nature Neuroscience.

[6]  J. Jeon,et al.  Transglutaminase 2 mediates transcriptional regulation through BAF250a polyamination , 2021, Genes & Genomics.

[7]  V. Rothhammer,et al.  Protective Functions of Reactive Astrocytes Following Central Nervous System Insult , 2020, Frontiers in Immunology.

[8]  M. Selzer,et al.  Advances in the Signaling Pathways Downstream of Glial-Scar Axon Growth Inhibitors , 2020, Frontiers in Cellular Neuroscience.

[9]  F. C. Bennett,et al.  INGE GRUNDKE-IQBAL AWARD FOR ALZHEIMER’S RESEARCH: NEUROTOXIC REACTIVE ASTROCYTES ARE INDUCED BY ACTIVATED MICROGLIA , 2019, Alzheimer's & Dementia.

[10]  R. Nuydens,et al.  High-throughput microscopy exposes a pharmacological window in which dual leucine zipper kinase inhibition preserves neuronal network connectivity , 2019, Acta Neuropathologica Communications.

[11]  H. Kitagawa,et al.  Glycan sulfation patterns define autophagy flux at axon tip via PTPRσ-cortactin axis , 2019, Nature Chemical Biology.

[12]  J. Lippincott-Schwartz,et al.  Neuron-Astrocyte Metabolic Coupling Protects against Activity-Induced Fatty Acid Toxicity , 2019, Cell.

[13]  G. Johnson,et al.  Nuclear transglutaminase 2 directly regulates expression of cathepsin S in rat cortical neurons , 2018, The European journal of neuroscience.

[14]  G. Johnson,et al.  Depletion of astrocytic transglutaminase 2 improves injury outcomes , 2018, Molecular and Cellular Neuroscience.

[15]  S. Saccani,et al.  Zbtb7a is a transducer for the control of promoter accessibility by NF-kappa B and multiple other transcription factors , 2018, PLoS biology.

[16]  G. Johnson,et al.  Transglutaminase 2: Friend or foe? The discordant role in neurons and astrocytes , 2018, Journal of neuroscience research.

[17]  H. M. Geller,et al.  Effect of chondroitin sulfate proteoglycans on neuronal cell adhesion, spreading and neurite growth in culture , 2018, Neural regeneration research.

[18]  G. Johnson,et al.  Subcellular localization patterns of transglutaminase 2 in astrocytes and neurons are differentially altered by hypoxia , 2017, Neuroreport.

[19]  G. Johnson,et al.  Transglutaminase 2 modulation of NF-κB signaling in astrocytes is independent of its ability to mediate astrocytic viability in ischemic injury , 2017, Brain Research.

[20]  M. Sofroniew,et al.  Cell biology of spinal cord injury and repair. , 2017, The Journal of clinical investigation.

[21]  H. Mansvelder,et al.  Astrocyte lipid metabolism is critical for synapse development and function in vivo , 2017, Glia.

[22]  S. Kojima,et al.  Transglutaminase 2 has opposing roles in the regulation of cellular functions as well as cell growth and death , 2016, Cell Death and Disease.

[23]  J. Harrill,et al.  Ontogeny of biochemical, morphological and functional parameters of synaptogenesis in primary cultures of rat hippocampal and cortical neurons , 2015, Molecular Brain.

[24]  N. J. Allen,et al.  Astrocytes Control Synapse Formation, Function, and Elimination. , 2015, Cold Spring Harbor perspectives in biology.

[25]  Y. Ao,et al.  Heterogeneity of reactive astrocytes , 2014, Neuroscience Letters.

[26]  R. Eckert,et al.  Transglutaminase regulation of cell function. , 2014, Physiological reviews.

[27]  P. D. Di Cesare,et al.  The Oncogene LRF Stimulates Proliferation of Mesenchymal Stem Cells and Inhibits Their Chondrogenic Differentiation , 2013, Cartilage.

[28]  N. Chabot,et al.  Transglutaminase and Polyamination of Tubulin: Posttranslational Modification for Stabilizing Axonal Microtubules , 2013, Neuron.

[29]  T. Yamashita,et al.  Chondroitin Sulfate Proteoglycans Down-regulate Spine Formation in Cortical Neurons by Targeting Tropomyosin-related Kinase B (TrkB) Protein , 2012, The Journal of Biological Chemistry.

[30]  G. Johnson,et al.  Complete transglutaminase 2 ablation results in reduced stroke volumes and astrocytes that exhibit increased survival in response to ischemia , 2012, Neurobiology of Disease.

[31]  G. Johnson,et al.  Transglutaminase 2: a molecular Swiss army knife. , 2012, Biochimica et biophysica acta.

[32]  J. Bol,et al.  Appearance of Tissue Transglutaminase in Astrocytes in Multiple Sclerosis Lesions: A Role in Cell Adhesion and Migration? , 2011, Brain pathology.

[33]  P. Dolan,et al.  Transglutaminase 2 protects against ischemic stroke , 2010, Neurobiology of Disease.

[34]  M. Sofroniew,et al.  Astrocytes: biology and pathology , 2009, Acta Neuropathologica.

[35]  M. Sofroniew Molecular dissection of reactive astrogliosis and glial scar formation , 2009, Trends in Neurosciences.

[36]  B. Barres The Mystery and Magic of Glia: A Perspective on Their Roles in Health and Disease , 2008, Neuron.

[37]  T. Osborne,et al.  Proto-oncogene FBI-1 (Pokemon) and SREBP-1 Synergistically Activate Transcription of Fatty-acid Synthase Gene (FASN)* , 2008, Journal of Biological Chemistry.

[38]  Fei Tan,et al.  Chondroitin-4-sulfation negatively regulates axonal guidance and growth , 2008, Journal of Cell Science.

[39]  M. Laudes,et al.  Transcription factor FBI-1 acts as a dual regulator in adipogenesis by coordinated regulation of cyclin-A and E2F-4 , 2008, Journal of Molecular Medicine.

[40]  L. Lorand,et al.  Mechanism of allosteric regulation of transglutaminase 2 by GTP , 2006, Proceedings of the National Academy of Sciences.

[41]  Zhigang He,et al.  Glial inhibition of CNS axon regeneration , 2006, Nature Reviews Neuroscience.

[42]  H. Müller,et al.  Collagen matrix in spinal cord injury. , 2006, Journal of neurotrauma.

[43]  George M. Smith,et al.  Growth factor and cytokine regulation of chondroitin sulfate proteoglycans by astrocytes , 2005, Glia.

[44]  G. Calapai,et al.  Transglutaminase activity and transglutaminase mRNA transcripts in gerbil brain ischemia , 2004, Neuroscience Letters.

[45]  Rita Casadio,et al.  Transglutaminases: nature's biological glues. , 2002, The Biochemical journal.

[46]  Sui Huang,et al.  FBI-1 can stimulate HIV-1 Tat activity and is targeted to a novel subnuclear domain that includes the Tat-P-TEFb-containing nuclear speckles. , 2002, Molecular biology of the cell.

[47]  S. Akimov,et al.  Cell-surface transglutaminase promotes fibronectin assembly via interaction with the gelatin-binding domain of fibronectin: a role in TGFbeta-dependent matrix deposition. , 2001, Journal of cell science.

[48]  K. Mehta,et al.  Tissue transglutaminase: an enzyme with a split personality. , 1999, The international journal of biochemistry & cell biology.

[49]  G. Johnson,et al.  Distinct Nuclear Localization and Activity of Tissue Transglutaminase* , 1998, The Journal of Biological Chemistry.

[50]  Stacey P. Memberg,et al.  Regeneration of adult axons in white matter tracts of the central nervous system , 1997, Nature.

[51]  D. Reinberg,et al.  Histone Deacetylases and SAP18, a Novel Polypeptide, Are Components of a Human Sin3 Complex , 1997, Cell.

[52]  O. Eizenberg,et al.  Expression of GTP-dependent and GTP-independent Tissue-type Transglutaminase in Cytokine-treated Rat Brain Astrocytes* , 1997, The Journal of Biological Chemistry.

[53]  S. Hoffman,et al.  Receptor-mediated adhesive and anti-adhesive functions of chondroitin sulfate proteoglycan preparations from embryonic chicken brain. , 1995, Journal of cell science.

[54]  D. Snow,et al.  Neurite outgrowth on a step gradient of chondroitin sulfate proteoglycan (CS-PG). , 1992, Journal of neurobiology.

[55]  R. Ientile,et al.  Transglutaminase 2 and neuroinflammation , 2014, Amino Acids.

[56]  G. Johnson,et al.  Transglutaminase 2 facilitates or ameliorates HIF signaling and ischemic cell death depending on its conformation and localization. , 2013, Biochimica et biophysica acta.

[57]  G. Johnson,et al.  The FASEB Journal • Research Communication Transglutaminase 2 protects against ischemic insult, interacts with HIF1�, and attenuates HIF1 signaling , 2022 .