Mono- and Biallelic Inactivation of Huntingtin Gene in Patient-Specific Induced Pluripotent Stem Cells Reveal HTT Roles in Striatal Development and Neuronal Functions

Overview

Journal J Huntingtons Dis

Publisher Sage Publications

Date 2024 Mar 1

PMID 38427495

Authors

Morgane Louessard

Michel Cailleret

Margot Jarrige

Julie Bigarreau

Sophie Lenoir

Noelle Dufour

Maria Rey

Frederic Saudou

Nicole Deglon

Anselme L Perrier

Affiliations

Soon will be listed here.

Abstract

Background: Mutations in the Huntingtin (HTT) gene cause Huntington's disease (HD), a neurodegenerative disorder. As a scaffold protein, HTT is involved in numerous cellular functions, but its normal and pathogenic functions during human forebrain development are poorly understood.

Objective: To investigate the developmental component of HD, with a specific emphasis on understanding the functions of wild-type and mutant HTT alleles during forebrain neuron development in individuals carrying HD mutations.

Methods: We used CRISPR/Cas9 gene-editing technology to disrupt the ATG region of the HTT gene via non-homologous end joining to produce mono- or biallelic HTT knock-out human induced pluripotent stem cell (iPSC) clones.

Results: We showed that the loss of wild-type, mutant, or both HTT isoforms does not affect the pluripotency of iPSCs or their transition into neural cells. However, we observed that HTT loss causes division impairments in forebrain neuro-epithelial cells and alters maturation of striatal projection neurons (SPNs) particularly in the acquisition of DARPP32 expression, a key functional marker of SPNs. Finally, young post-mitotic neurons derived from HTT-/- human iPSCs display cellular dysfunctions observed in adult HD neurons.

Conclusions: We described a novel collection of isogenic clones with mono- and biallelic HTT inactivation that complement existing HD-hiPSC isogenic series to explore HTT functions and test therapeutic strategies in particular HTT-lowering drugs. Characterizing neural and neuronal derivatives from human iPSCs of this collection, we show evidence that HTT loss or mutation has impacts on neuro-epithelial and striatal neurons maturation, and on basal DNA damage and BDNF axonal transport in post-mitotic neurons.

References

Madabhushi R, Pan L, Tsai L . DNA damage and its links to neurodegeneration. Neuron. 2014; 83(2):266-282. PMC: 5564444. DOI: 10.1016/j.neuron.2014.06.034. View

Kaemmerer W, Grondin R . The effects of huntingtin-lowering: what do we know so far?. Degener Neurol Neuromuscul Dis. 2019; 9:3-17. PMC: 6413743. DOI: 10.2147/DNND.S163808. View

Jung R, Lee Y, Barker D, Correia K, Shin B, Loupe J . Mutations causing Lopes-Maciel-Rodan syndrome are huntingtin hypomorphs. Hum Mol Genet. 2021; 30(3-4):135-148. PMC: 8248964. DOI: 10.1093/hmg/ddaa283. View

Zhang S, Feany M, Saraswati S, Littleton J, Perrimon N . Inactivation of Drosophila Huntingtin affects long-term adult functioning and the pathogenesis of a Huntington's disease model. Dis Model Mech. 2009; 2(5-6):247-66. PMC: 2675792. DOI: 10.1242/dmm.000653. View

Feyeux M, Bourgois-Rocha F, Redfern A, Giles P, Lefort N, Aubert S . Early transcriptional changes linked to naturally occurring Huntington's disease mutations in neural derivatives of human embryonic stem cells. Hum Mol Genet. 2012; 21(17):3883-95. DOI: 10.1093/hmg/dds216. View

Van Raamsdonk J, Pearson J, Rogers D, Bissada N, Vogl A, Hayden M . Loss of wild-type huntingtin influences motor dysfunction and survival in the YAC128 mouse model of Huntington disease. Hum Mol Genet. 2005; 14(10):1379-92. DOI: 10.1093/hmg/ddi147. View

Gauthier L, Charrin B, Borrell-Pages M, Dompierre J, Rangone H, Cordelieres F . Huntingtin controls neurotrophic support and survival of neurons by enhancing BDNF vesicular transport along microtubules. Cell. 2004; 118(1):127-38. DOI: 10.1016/j.cell.2004.06.018. View

Arber C, Precious S, Cambray S, Risner-Janiczek J, Kelly C, Noakes Z . Activin A directs striatal projection neuron differentiation of human pluripotent stem cells. Development. 2015; 142(7):1375-86. PMC: 4378247. DOI: 10.1242/dev.117093. View

Ooi J, Langley S, Xu X, Utami K, Sim B, Huang Y . Unbiased Profiling of Isogenic Huntington Disease hPSC-Derived CNS and Peripheral Cells Reveals Strong Cell-Type Specificity of CAG Length Effects. Cell Rep. 2019; 26(9):2494-2508.e7. DOI: 10.1016/j.celrep.2019.02.008. View

10.

. Identification of Genetic Factors that Modify Clinical Onset of Huntington's Disease. Cell. 2015; 162(3):516-26. PMC: 4524551. DOI: 10.1016/j.cell.2015.07.003. View

11.

Molina-Calavita M, Barnat M, Elias S, Aparicio E, Piel M, Humbert S . Mutant huntingtin affects cortical progenitor cell division and development of the mouse neocortex. J Neurosci. 2014; 34(30):10034-40. PMC: 6608303. DOI: 10.1523/JNEUROSCI.0715-14.2014. View

12.

Gribaudo S, Bousset L, Courte J, Melki R, Peyrin J, Perrier A . Propagation of Distinct α-Synuclein Strains Within Human Reconstructed Neuronal Network and Associated Neuronal Dysfunctions. Methods Mol Biol. 2022; 2551:357-378. DOI: 10.1007/978-1-0716-2597-2_24. View

13.

Virlogeux A, Moutaux E, Christaller W, Genoux A, Bruyere J, Fino E . Reconstituting Corticostriatal Network on-a-Chip Reveals the Contribution of the Presynaptic Compartment to Huntington's Disease. Cell Rep. 2018; 22(1):110-122. DOI: 10.1016/j.celrep.2017.12.013. View

14.

Reiner A, Albin R, Anderson K, DAMATO C, Penney J, Young A . Differential loss of striatal projection neurons in Huntington disease. Proc Natl Acad Sci U S A. 1988; 85(15):5733-7. PMC: 281835. DOI: 10.1073/pnas.85.15.5733. View

15.

Drouet V, Ruiz M, Zala D, Feyeux M, Auregan G, Cambon K . Allele-specific silencing of mutant huntingtin in rodent brain and human stem cells. PLoS One. 2014; 9(6):e99341. PMC: 4057216. DOI: 10.1371/journal.pone.0099341. View

16.

Enokido Y, Tamura T, Ito H, Arumughan A, Komuro A, Shiwaku H . Mutant huntingtin impairs Ku70-mediated DNA repair. J Cell Biol. 2010; 189(3):425-43. PMC: 2867301. DOI: 10.1083/jcb.200905138. View

17.

Burrus C, McKinstry S, Kim N, Ozlu M, Santoki A, Fang F . Striatal Projection Neurons Require Huntingtin for Synaptic Connectivity and Survival. Cell Rep. 2020; 30(3):642-657.e6. PMC: 7025500. DOI: 10.1016/j.celrep.2019.12.069. View

18.

Nasir J, Floresco S, OKusky J, Diewert V, Richman J, Zeisler J . Targeted disruption of the Huntington's disease gene results in embryonic lethality and behavioral and morphological changes in heterozygotes. Cell. 1995; 81(5):811-23. DOI: 10.1016/0092-8674(95)90542-1. View

19.

Hensman Moss D, Pardinas A, Langbehn D, Lo K, Leavitt B, Roos R . Identification of genetic variants associated with Huntington's disease progression: a genome-wide association study. Lancet Neurol. 2017; 16(9):701-711. DOI: 10.1016/S1474-4422(17)30161-8. View

20.

Reiner A, Del Mar N, Meade C, Yang H, Dragatsis I, Zeitlin S . Neurons lacking huntingtin differentially colonize brain and survive in chimeric mice. J Neurosci. 2001; 21(19):7608-19. PMC: 6762912. View