MARIA GIOVANNA GARONE

PhD Graduate

PhD program:: XXXIV


advisor: Alessandro Rosa
co-supervisor: Alessandro Rosa

Thesis title: RNA-binding protein network alteration causes aberrant axon branching and growth phenotypes in FUS-ALS mutant motor neurons.

Mutations in RNA-binding proteins (RBPs) have been genetically associated with the motoneuron disease Amyotrophic Lateral Sclerosis (ALS), suggesting a causative link between dysregulation of RNA metabolism and this neurodegenerative disease. However, the lack of mechanistic insights has hampered the development of therapeutic approaches. In this regard, my work aimed to evaluate the toxic functions gained by the ALS-linked gene FUS, with a specific mechanistic focus on the aberrant crosstalk between mutant FUS and other RBPs. To achieve this goal, a novel protocol for obtaining functional spinal and cranial motor neurons (MNs) from human induced pluripotent stem cells (iPSCs) based on ectopic transcription factor modules was developed. By programming rapid cell differentiation into a pure population with high efficiency and reproducibility, this method overcomes significant issues of converting iPSCs to specific disease-relevant cell types (Garone et al., 2019; Garone e Rosa, 2021). iPSC-derived MNs were used to study the effects of ALS mutant FUS on the human motoneuronal proteome. This analysis revealed that, in FUS mutant MNs, proteins involved in catabolic processes and oxidation-reduction are upregulated, whereas cytoskeletal proteins and factors driving neuron projection are downregulated. Interestingly, proteome alteration does not correlate with transcriptome changes. Rather, a strong correlation with selective binding was observed by mutant FUS to mRNAs in their 3’UTR (Garone et al., 2020). To gain mechanistic insights, a relevant FUS target, the neural RBP HuD (encoded by the ELAVL4 gene), was investigated. In both iPSC-derived MNs and FUS mouse model, mutant FUS leads to upregulation of HuD protein levels through competition for HuD 3’UTR binding with FMRP, an RBP encoded by the Fragile-X gene FMR1. Indeed, FMRP was found as a novel repressor of HuD translation, and mutant FUS disrupts this function. In turn, increased HuD levels overly stabilize the transcript of two key targets, NRN1 and GAP43. Both factors are involved in axon growth and, accordingly, axon branching and growth upon injury were increased in FUS mutant MNs. Notably, such phenotypes could be rescued by dampening NRN1 levels (Garone et al., 2021). Since similar axonal phenotypes have been previously described in SOD1 and TDP-43 mutant models, aberrant axonal growth and branching might represent broad early events in the pathogenesis of ALS. Thus, based on this work, novel therapeutic approaches targeting HuD and/or NRN1 and GAP43 might be pursued for ALS in the future. Moreover, evidence of cross-regulation between HuD and FMRP, two important neural RBPs playing multiple roles in nervous system development and neurological diseases, has implications that may extend beyond the ALS field.

Research products

11573/1668400 - 2023 - Digital color-coded molecular barcoding reveals dysregulation of common FUS and FMRP targets in soma and neurites of ALS mutant motoneurons
Garone, Maria Giovanna; Salerno, Debora; Rosa, Alessandro - 01a Articolo in rivista
paper: CELL DEATH DISCOVERY ([London]: Nature Publishing Group, 2015-) pp. - - issn: 2058-7716 - wos: WOS:000920816800001 (1) - scopus: 2-s2.0-85146766524 (2)

11573/1692082 - 2023 - The long noncoding RNA nHOTAIRM1 is necessary for differentiation and activity of iPSC-derived spinal motor neurons
Tollis, P.; Vitiello, E.; Migliaccio, F.; D'ambra, E.; Rocchegiani, A.; Garone, M. G.; Bozzoni, I.; Rosa, A.; Carissimo, A.; Laneve, P.; Caffarelli, E. - 01a Articolo in rivista
paper: CELL DEATH & DISEASE (Nature Group) pp. - - issn: 2041-4889 - wos: WOS:001101872900003 (2) - scopus: 2-s2.0-85176472807 (2)

11573/1645416 - 2022 - Culture of Human iPSC-Derived Motoneurons in Compartmentalized Microfluidic Devices and Quantitative Assays for Studying Axonal Phenotypes
Garone, M. G.; D'antoni, C.; Rosa, A. - 02a Capitolo o Articolo
book: Stem Cell Assays: Methods and Protocols - (978-1-0716-1978-0; 978-1-0716-1979-7)

11573/1662787 - 2022 - Emerging roles for the RNA-Binding protein HuD (ELAVL4) in nervous system diseases
Silvestri, Beatrice; Mochi, Michela; Garone, Maria Giovanna; Rosa, Alessandro - 01g Articolo di rassegna (Review)
paper: INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES (Basel: MDPI Center) pp. - - issn: 1422-0067 - wos: WOS:000896194600001 (6) - scopus: 2-s2.0-85143789148 (8)

11573/1564033 - 2021 - FUS-ALS mutants alter FMRP phase separation equilibrium and impair protein translation
Birsa, N.; Ule, A. M.; Garone, M. G.; Tsang, B.; Mattedi, F.; Andrew Chong, P.; Humphrey, J.; Jarvis, S.; Pisiren, M.; Wilkins, O. G.; Nosella, M. L.; Devoy, A.; Bodo, C.; De La Fuente, R. F.; Fisher, E. M. C.; Rosa, A.; Viero, G.; Forman-Kay, J. D.; Schiavo, G.; Fratta, P. - 01a Articolo in rivista
paper: SCIENCE ADVANCES (Washington, DC : American Association for the Advancement of Science, 2015-) pp. 1-16 - issn: 2375-2548 - wos: WOS:000675849000015 (30) - scopus: 2-s2.0-85110283030 (30)

11573/1567941 - 2021 - ALS-related FUS mutations alter axon growth in motoneurons and affect HuD/ELAVL4 and FMRP activity
Garone, M. G.; Birsa, N.; Rosito, M.; Salaris, F.; Mochi, M.; De Turris, V.; Nair, R. R.; Cunningham, T. J.; Fisher, E. M. C.; Morlando, M.; Fratta, P.; Rosa, A. - 01a Articolo in rivista
paper: COMMUNICATIONS BIOLOGY ([London]: Springer Nature) pp. 1025- - issn: 2399-3642 - wos: WOS:000692407000006 (19) - scopus: 2-s2.0-85114160672 (23)

11573/1631026 - 2021 - PiggyBac vectors in pluripotent stem cell research and applications
Garone, M. G.; Rosa, A. - 02a Capitolo o Articolo
book: Methods in iPSC Technology - (9780323857666)

11573/1549404 - 2021 - Small heat-shock protein HSPB3 promotes myogenesis by regulating the lamin B receptor
Tiago, T.; Hummel, B.; Morelli, F. F.; Basile, V.; Vinet, J.; Galli, V.; Mediani, L.; Antoniani, F.; Pomella, S.; Cassandri, M.; Garone, M. G.; Silvestri, B.; Cimino, M.; Cenacchi, G.; Costa, R.; Mouly, V.; Poser, I.; Yeger-Lotem, E.; Rosa, A.; Alberti, S.; Rota, R.; Ben-Zvi, A.; Sawarkar, R.; Carra, S. - 01a Articolo in rivista
paper: CELL DEATH & DISEASE (Nature Group) pp. 1-19 - issn: 2041-4889 - wos: WOS:000656257000003 (12) - scopus: 2-s2.0-85105454628 (15)

11573/1430664 - 2020 - Proteomics analysis of FUS mutant human motoneurons reveals altered regulation of cytoskeleton and other ALS-linked proteins via 3′UTR binding
Garone, Maria Giovanna; Alfano, Vincenzo; Salvatori, Beatrice; Braccia, Clarissa; Peruzzi, Giovanna; Colantoni, Alessio; Bozzoni, Irene; Armirotti, Andrea; Rosa, Alessandro - 01a Articolo in rivista
paper: SCIENTIFIC REPORTS (London: Springer Nature London: Nature Publishing Group) pp. - - issn: 2045-2322 - wos: WOS:000550057200140 (12) - scopus: 2-s2.0-85087975715 (15)

11573/1405967 - 2020 - FUS ALS-causative mutations impair FUS autoregulation and splicing factor networks through intron retention
Humphrey, Jack; Birsa, Nicol; Milioto, Carmelo; Mclaughlin, Martha; Ule, Agnieszka M; Robaldo, David; Eberle, Andrea B; Kräuchi, Rahel; Bentham, Matthew; Brown, Anna-Leigh; Jarvis, Seth; Bodo, Cristian; Garone, Maria G; Devoy, Anny; Soraru, Gianni; Rosa, Alessandro; Bozzoni, Irene; Fisher, Elizabeth M C; Mühlemann, Oliver; Schiavo, Giampietro; Ruepp, Marc-David; Isaacs, Adrian M; Plagnol, Vincent; Fratta, Pietro - 01a Articolo in rivista
paper: NUCLEIC ACIDS RESEARCH (Oxford University Press:Journals Department, Great Clarendon Street, Oxford OX2 6DP United Kingdom:011 44 1865 556767, EMAIL: jnlorders@oup.co.uk, INTERNET: http://www.oup.co.uk, Fax: 011 44 1865 267485 Editore precedente: Information Retrieval ltd., London) pp. - - issn: 0305-1048 - wos: WOS:000574288800041 (56) - scopus: 2-s2.0-85088208506 (60)

11573/1429819 - 2020 - HOTAIRM1 regulates neuronal differentiation by modulating NEUROGENIN 2 and the downstream neurogenic cascade
Rea, Jessica; Menci, Valentina; Tollis, Paolo; Santini, Tiziana; Armaos, Alexandros; Garone, Maria Giovanna; Iberite, Federica; Cipriano, Andrea; Tartaglia, Gian Gaetano; Rosa, Alessandro; Ballarino, Monica; Laneve, Pietro; Caffarelli, Elisa - 01a Articolo in rivista
paper: CELL DEATH & DISEASE (Nature Group) pp. - - issn: 2041-4889 - wos: WOS:000553100400002 (31) - scopus: 2-s2.0-85087954274 (27)

11573/1297233 - 2019 - Mutant FUS and ELAVL4 (HuD) aberrant crosstalk in Amyotrophic Lateral Sclerosis
De Santis, R.; Alfano, V.; De Turris, V.; Colantoni, A.; Santini, L.; Garone, M. G.; Antonacci, G.; Peruzzi, G.; Sudria-Lopez, E.; Wyler, E.; Anink, J. J.; Aronica, E.; Landthaler, M.; Pasterkamp, R. J.; Bozzoni, I.; Rosa, A. - 01a Articolo in rivista
paper: CELL REPORTS (Elsevier B.V. Cambridge MA: Cell Press) pp. 3818-3831 - issn: 2211-1247 - wos: WOS:000472715000011 (40) - scopus: 2-s2.0-85067263880 (42)

11573/1276468 - 2019 - Conversion of Human Induced Pluripotent Stem Cells (iPSCs) into Functional Spinal and Cranial Motor Neurons Using PiggyBac Vectors
Garone, Maria G; De Turris, Valeria; Soloperto, Alessandro; Brighi, Carlo; De Santis, Riccardo; Pagani, Francesca; Di Angelantonio, Silvia; Rosa, Alessandro - 01a Articolo in rivista
paper: JOURNAL OF VISUALIZED EXPERIMENTS ([Boston, Mass. : MYJoVE Corp., 2006]-) pp. - - issn: 1940-087X - wos: WOS:000469977600054 (13) - scopus: 2-s2.0-85066930236 (18)

11573/1345007 - 2019 - HOTAIRM1 regulates neuronal differentiation by controlling NEUROGENIN 2 and the downstream neurogenic cascade
Rea, Jessica; Menci, Valentina; Tollis, Paolo; Santini, Tiziana; Alexandros, Armaos; Garone, Maria Giovanna; Iberite, Federica; Cipriano, Andrea; Tartaglia, Gian Gaetano; Rosa, Alessandro; Ballarino, Monica; Laneve, Pietro; Caffarelli, Elisa - 04f Poster
conference: EMBO | EMBL Symposium: The Non-Coding Genome (EMBL Heidelberg, Germany)
book: EMBO | EMBL Symposium: The Non-Coding Genome - ABSTRACT BOOK - ()

11573/1109389 - 2018 - Direct conversion of human pluripotent stem cells into cranial motor neurons using a piggyBac vector
De Santis, Riccardo; Garone, Maria Giovanna; Pagani, Francesca; De Turris, Valeria; Di Angelantonio, Silvia; Rosa, Alessandro - 01a Articolo in rivista
paper: STEM CELL RESEARCH (Amsterdam : Elsevier) pp. 189-196 - issn: 1873-5061 - wos: WOS:000434978900033 (29) - scopus: 2-s2.0-85046443648 (33)

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