Nuclear-architecture defects have been shown to correlate with the manifestation of a number of human diseases as well as ageing1,2,3,4. It is therefore plausible that diseases whose manifestations correlate with ageing might be connected to the appearance of nuclear aberrations over time. We decided to evaluate nuclear organization in the context of ageing-associated disorders by focusing on a leucine-rich repeat kinase 2 (LRRK2) dominant mutation (G2019S; glycine-to-serine substitution at amino acid 2019), which is associated with familial and sporadic Parkinson’s disease as well as impairment of adult neurogenesis in mice5. Here we report on the generation of induced pluripotent stem cells (iPSCs) derived from Parkinson’s disease patients and the implications of LRRK2(G2019S) mutation in human neural-stem-cell (NSC) populations. Mutant NSCs showed increased susceptibility to proteasomal stress as well as passage-dependent deficiencies in nuclear-envelope organization, clonal expansion and neuronal differentiation. Disease phenotypes were rescued by targeted correction of the LRRK2(G2019S) mutation with its wild-type counterpart in Parkinson’s disease iPSCs and were recapitulated after targeted knock-in of the LRRK2(G2019S) mutation in human embryonic stem cells. Analysis of human brain tissue showed nuclear-envelope impairment in clinically diagnosed Parkinson’s disease patients. Together, our results identify the nucleus as a previously unknown cellular organelle in Parkinson’s disease pathology and may help to open new avenues for Parkinson’s disease diagnoses as well as for the potential development of therapeutics targeting this fundamental cell structure.
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We would like to thank K. Mitani, P. Ng, A. Lieber, Y. Imai, M. A. Miyawaki, Filocamo, S. Goldwurm, Telethon Genetic Biobank Network for providing constructs and cells (the fibroblast samples were obtained from the “Cell Line and DNA Biobank from patients affected by Genetic Diseases” (G. Gaslini Institute)-Telethon Genetic Biobank Network (project no. GTB07001)); Neurological Tissue Bank of the Biobank-Hospital Clínic-IDIBAPS for providing human brain tissue; F. Gage, M. Hetzer, J. Yao, Y. Mu, D. Yu, E. Gelpí, X. M. Wang, X. Wang, G. Bai and Z. J. Liu for helpful discussions; M. Joens and J. Fitzpatrick of the Waitt Advanced Biophotonics Core Facility for performing TEM analysis; M. Marti for imaging, teratoma and karyotyping analysis; F. Osakada for statistics analysis; and M. Schwarz, P. Schwarz and L. Laricchia-Robbio for administrative help. G.-H.L. is supported by the Thousand Young Talents program of China, the National Laboratory of Biomacromolecules, the Strategic Priority Research Program of the Chinese Academy of Sciences, the National Natural Science Foundation of China (NSFC) (81271266 and 31222039), and the Beijing Municipal Natural Science Foundation. J.Q. was partly supported by an AFAR/Ellison Medical Foundation postdoctoral fellowship. K.S. was partly supported by a Uehara Memorial Foundation research fellowship. E.N. was partly supported by an F.M. Kirby Foundation postdoctoral fellowship. X.X. is supported by NSFC (31201111). B.R. was supported by a US National Institute of Health (NIH) grant (ES017166) and the Ludwig Institute for Cancer Research. J.Y. was supported by an NIH grant (P41 RR011823). J.C.I.B. was supported by grants from the Glenn Foundation, G. Harold and Leila Y. Mathers Charitable Foundation, Sanofi, the California Institute of Regenerative Medicine, the Ellison Medical Foundation, the Helmsley Charitable Trust, ERA-Net Neuron, MINECO and Fundacion Cellex.
The authors declare no competing financial interests.
This file contains Supplementary Methods, Supplementary Figures 1-18 and Supplementary references. (PDF 3162 kb)
This file contains Supplementary Tables 1-5. (XLS 135 kb)
The In-1 mediated restoration of cellular morphology in late passage LRRK2 G2019S NSCs. 5 mM In-1 was added to passage 18 ipsNSCs-LK2(GS/GS), and then the cells were cultured for 5 days. Cells were imaged every 10 min for the 5 day duration. (MOV 18568 kb)
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Liu, GH., Qu, J., Suzuki, K. et al. Progressive degeneration of human neural stem cells caused by pathogenic LRRK2. Nature 491, 603–607 (2012). https://doi.org/10.1038/nature11557
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