CRISPR/Cas9 and piggyBac-mediated footprint-free LRRK2-G2019S knock-in reveals neuronal complexity phenotypes and α-Synuclein modulation in dopaminergic neurons
The p.G2019S mutation of the leucine-rich repeat kinase 2 (LRRK2) has been identified as the most prevalent genetic cause of familial and sporadic Parkinson's disease (PD). The Cre-LoxP recombination system has been used to correct the LRRK2-G2019S mutation in patient derived human induced plur...
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| Format: | Article |
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Elsevier
2017-10-01
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| Series: | Stem Cell Research |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S187350611730168X |
| _version_ | 1811198827775918080 |
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| author | Xiaobing Qing Jonas Walter Javier Jarazo Jonathan Arias-Fuenzalida Anna-Lena Hillje Jens C. Schwamborn |
| author_facet | Xiaobing Qing Jonas Walter Javier Jarazo Jonathan Arias-Fuenzalida Anna-Lena Hillje Jens C. Schwamborn |
| author_sort | Xiaobing Qing |
| collection | DOAJ |
| description | The p.G2019S mutation of the leucine-rich repeat kinase 2 (LRRK2) has been identified as the most prevalent genetic cause of familial and sporadic Parkinson's disease (PD). The Cre-LoxP recombination system has been used to correct the LRRK2-G2019S mutation in patient derived human induced pluripotent stem cells (hiPSCs) in order to generate isogenic controls. However, the remaining LoxP site can influence gene expression. In this study, we report the generation of a footprint-free LRRK2-G2019S isogenic hiPS cell line edited with the CRISPR/Cas9 and piggyBac technologies. We observed that the percentage of Tyrosine Hydroxylase (TH) positive neurons with a total neurite length of >2000 μm was significantly reduced in LRRK2-G2019S dopaminergic (DA) neurons. The average branch number in LRRK2-G2019S DA neurons was also decreased. In addition, we have shown that in vitro TH positive neurons with a total neurite length of >2000 μm were positive for Serine 129 phosphorylated (S129P) alpha-Synuclein (αS) and we hypothesize that S129P-αS plays a role in the maintenance or formation of long neurites. In summary, our footprint-free LRRK2-G2019S isogenic cell lines allow standardized, genetic background independent, in vitro PD modeling and provide new insights into the role of LRRK2-G2019S and S129P-αS in the pathogenesis of PD. |
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| institution | Directory Open Access Journal |
| issn | 1873-5061 1876-7753 |
| language | English |
| last_indexed | 2024-04-12T01:37:54Z |
| publishDate | 2017-10-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Stem Cell Research |
| spelling | doaj.art-fd5abaaaa9a841d087806577035851e52022-12-22T03:53:17ZengElsevierStem Cell Research1873-50611876-77532017-10-0124C445010.1016/j.scr.2017.08.013CRISPR/Cas9 and piggyBac-mediated footprint-free LRRK2-G2019S knock-in reveals neuronal complexity phenotypes and α-Synuclein modulation in dopaminergic neuronsXiaobing QingJonas WalterJavier JarazoJonathan Arias-FuenzalidaAnna-Lena HilljeJens C. SchwambornThe p.G2019S mutation of the leucine-rich repeat kinase 2 (LRRK2) has been identified as the most prevalent genetic cause of familial and sporadic Parkinson's disease (PD). The Cre-LoxP recombination system has been used to correct the LRRK2-G2019S mutation in patient derived human induced pluripotent stem cells (hiPSCs) in order to generate isogenic controls. However, the remaining LoxP site can influence gene expression. In this study, we report the generation of a footprint-free LRRK2-G2019S isogenic hiPS cell line edited with the CRISPR/Cas9 and piggyBac technologies. We observed that the percentage of Tyrosine Hydroxylase (TH) positive neurons with a total neurite length of >2000 μm was significantly reduced in LRRK2-G2019S dopaminergic (DA) neurons. The average branch number in LRRK2-G2019S DA neurons was also decreased. In addition, we have shown that in vitro TH positive neurons with a total neurite length of >2000 μm were positive for Serine 129 phosphorylated (S129P) alpha-Synuclein (αS) and we hypothesize that S129P-αS plays a role in the maintenance or formation of long neurites. In summary, our footprint-free LRRK2-G2019S isogenic cell lines allow standardized, genetic background independent, in vitro PD modeling and provide new insights into the role of LRRK2-G2019S and S129P-αS in the pathogenesis of PD.http://www.sciencedirect.com/science/article/pii/S187350611730168XLRRK2-G2019SCRISPR/Cas9piggyBacParkinson's diseasehiPSα-SynucleinSerine129 phosphorylated α-Synuclein |
| spellingShingle | Xiaobing Qing Jonas Walter Javier Jarazo Jonathan Arias-Fuenzalida Anna-Lena Hillje Jens C. Schwamborn CRISPR/Cas9 and piggyBac-mediated footprint-free LRRK2-G2019S knock-in reveals neuronal complexity phenotypes and α-Synuclein modulation in dopaminergic neurons Stem Cell Research LRRK2-G2019S CRISPR/Cas9 piggyBac Parkinson's disease hiPS α-Synuclein Serine129 phosphorylated α-Synuclein |
| title | CRISPR/Cas9 and piggyBac-mediated footprint-free LRRK2-G2019S knock-in reveals neuronal complexity phenotypes and α-Synuclein modulation in dopaminergic neurons |
| title_full | CRISPR/Cas9 and piggyBac-mediated footprint-free LRRK2-G2019S knock-in reveals neuronal complexity phenotypes and α-Synuclein modulation in dopaminergic neurons |
| title_fullStr | CRISPR/Cas9 and piggyBac-mediated footprint-free LRRK2-G2019S knock-in reveals neuronal complexity phenotypes and α-Synuclein modulation in dopaminergic neurons |
| title_full_unstemmed | CRISPR/Cas9 and piggyBac-mediated footprint-free LRRK2-G2019S knock-in reveals neuronal complexity phenotypes and α-Synuclein modulation in dopaminergic neurons |
| title_short | CRISPR/Cas9 and piggyBac-mediated footprint-free LRRK2-G2019S knock-in reveals neuronal complexity phenotypes and α-Synuclein modulation in dopaminergic neurons |
| title_sort | crispr cas9 and piggybac mediated footprint free lrrk2 g2019s knock in reveals neuronal complexity phenotypes and α synuclein modulation in dopaminergic neurons |
| topic | LRRK2-G2019S CRISPR/Cas9 piggyBac Parkinson's disease hiPS α-Synuclein Serine129 phosphorylated α-Synuclein |
| url | http://www.sciencedirect.com/science/article/pii/S187350611730168X |
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