Live cell in situ lysosomal GCase activity correlates to alpha-synuclein levels in human differentiated neurons with LRRK2 and GBA1 mutations
IntroductionHeterozygous mutations in GBA1, which encodes the lysosomal hydrolase glucocerebrosidase (GCase), are a common risk factor for the neurodegenerative movement disorder Parkinson's disease (PD). Consequently, therapeutic options targeting the GCase enzyme are in development. An import...
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Frontiers Media S.A.
2023-10-01
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| Series: | Frontiers in Cellular Neuroscience |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fncel.2023.1229213/full |
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| author | Adahir Labrador-Garrido Siying Zhong Laura Hughes Shikara Keshiya Woojin S. Kim Glenda M. Halliday Nicolas Dzamko |
| author_facet | Adahir Labrador-Garrido Siying Zhong Laura Hughes Shikara Keshiya Woojin S. Kim Glenda M. Halliday Nicolas Dzamko |
| author_sort | Adahir Labrador-Garrido |
| collection | DOAJ |
| description | IntroductionHeterozygous mutations in GBA1, which encodes the lysosomal hydrolase glucocerebrosidase (GCase), are a common risk factor for the neurodegenerative movement disorder Parkinson's disease (PD). Consequently, therapeutic options targeting the GCase enzyme are in development. An important aspect of this development is determining the effect of potential modifying compounds on GCase activity, which can be complicated by the different methods and substrate probes that are commonly employed for this purpose.MethodsIn this study, we employed the GCase substrate probe 5-(pentafluorobenzoylamino)fluorescein di-D-glucopyranoside (PFB-FDGlu) in combination with live cell imaging to measure GCase activity in situ in the lysosome.ResultsThe live cell assay was validated using the GCase inhibitor conduritol-B-epoxide and with GBA1 knockout neural cells and was then used to assess GCase activity in iPSC differentiated into neural stem cells and neurons that were obtained from idiopathic PD patients and PD patients with the LRRK2 G2019S and GBA N370S mutations, as well as controls (n = 4 per group). Heterogeneity in GCase activity was observed across all groups. However, a significant inverse correlation between GCase activity and levels of alpha-synuclein protein was observed.DiscussionThe live cell imaging assay for GCase activity could be useful for further understanding the role of GCase in PD and screening potential modifying compounds in differentiated human cell models. |
| first_indexed | 2024-03-11T18:16:31Z |
| format | Article |
| id | doaj.art-c89347aae76540ea9933273a0178d93c |
| institution | Directory Open Access Journal |
| issn | 1662-5102 |
| language | English |
| last_indexed | 2024-03-11T18:16:31Z |
| publishDate | 2023-10-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Cellular Neuroscience |
| spelling | doaj.art-c89347aae76540ea9933273a0178d93c2023-10-16T06:58:27ZengFrontiers Media S.A.Frontiers in Cellular Neuroscience1662-51022023-10-011710.3389/fncel.2023.12292131229213Live cell in situ lysosomal GCase activity correlates to alpha-synuclein levels in human differentiated neurons with LRRK2 and GBA1 mutationsAdahir Labrador-GarridoSiying ZhongLaura HughesShikara KeshiyaWoojin S. KimGlenda M. HallidayNicolas DzamkoIntroductionHeterozygous mutations in GBA1, which encodes the lysosomal hydrolase glucocerebrosidase (GCase), are a common risk factor for the neurodegenerative movement disorder Parkinson's disease (PD). Consequently, therapeutic options targeting the GCase enzyme are in development. An important aspect of this development is determining the effect of potential modifying compounds on GCase activity, which can be complicated by the different methods and substrate probes that are commonly employed for this purpose.MethodsIn this study, we employed the GCase substrate probe 5-(pentafluorobenzoylamino)fluorescein di-D-glucopyranoside (PFB-FDGlu) in combination with live cell imaging to measure GCase activity in situ in the lysosome.ResultsThe live cell assay was validated using the GCase inhibitor conduritol-B-epoxide and with GBA1 knockout neural cells and was then used to assess GCase activity in iPSC differentiated into neural stem cells and neurons that were obtained from idiopathic PD patients and PD patients with the LRRK2 G2019S and GBA N370S mutations, as well as controls (n = 4 per group). Heterogeneity in GCase activity was observed across all groups. However, a significant inverse correlation between GCase activity and levels of alpha-synuclein protein was observed.DiscussionThe live cell imaging assay for GCase activity could be useful for further understanding the role of GCase in PD and screening potential modifying compounds in differentiated human cell models.https://www.frontiersin.org/articles/10.3389/fncel.2023.1229213/fullParkinson's diseaseGCasealpha-synucleinpluripotent stem cellsLRRK2 |
| spellingShingle | Adahir Labrador-Garrido Siying Zhong Laura Hughes Shikara Keshiya Woojin S. Kim Glenda M. Halliday Nicolas Dzamko Live cell in situ lysosomal GCase activity correlates to alpha-synuclein levels in human differentiated neurons with LRRK2 and GBA1 mutations Frontiers in Cellular Neuroscience Parkinson's disease GCase alpha-synuclein pluripotent stem cells LRRK2 |
| title | Live cell in situ lysosomal GCase activity correlates to alpha-synuclein levels in human differentiated neurons with LRRK2 and GBA1 mutations |
| title_full | Live cell in situ lysosomal GCase activity correlates to alpha-synuclein levels in human differentiated neurons with LRRK2 and GBA1 mutations |
| title_fullStr | Live cell in situ lysosomal GCase activity correlates to alpha-synuclein levels in human differentiated neurons with LRRK2 and GBA1 mutations |
| title_full_unstemmed | Live cell in situ lysosomal GCase activity correlates to alpha-synuclein levels in human differentiated neurons with LRRK2 and GBA1 mutations |
| title_short | Live cell in situ lysosomal GCase activity correlates to alpha-synuclein levels in human differentiated neurons with LRRK2 and GBA1 mutations |
| title_sort | live cell in situ lysosomal gcase activity correlates to alpha synuclein levels in human differentiated neurons with lrrk2 and gba1 mutations |
| topic | Parkinson's disease GCase alpha-synuclein pluripotent stem cells LRRK2 |
| url | https://www.frontiersin.org/articles/10.3389/fncel.2023.1229213/full |
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