Autophagy and Lysosomal Functionality in CMT2B Fibroblasts Carrying the RAB7<sup>K126R</sup> Mutation
Charcot-Marie-Tooth type 2B (CMT2B) disease is a dominant axonal peripheral neuropathy caused by five mutations in the <i>RAB7A</i> gene. Autophagy and late endocytic trafficking were already characterized in CMT2B. Indeed, impairment of autophagy and an increase in lysosomal degradative...
| Main Authors: | , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2022-01-01
|
| Series: | Cells |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2073-4409/11/3/496 |
| _version_ | 1797488442395852800 |
|---|---|
| author | Roberta Romano Victoria Stefania Del Fiore Paola Saveri Ilaria Elena Palamà Chiara Pisciotta Davide Pareyson Cecilia Bucci Flora Guerra |
| author_facet | Roberta Romano Victoria Stefania Del Fiore Paola Saveri Ilaria Elena Palamà Chiara Pisciotta Davide Pareyson Cecilia Bucci Flora Guerra |
| author_sort | Roberta Romano |
| collection | DOAJ |
| description | Charcot-Marie-Tooth type 2B (CMT2B) disease is a dominant axonal peripheral neuropathy caused by five mutations in the <i>RAB7A</i> gene. Autophagy and late endocytic trafficking were already characterized in CMT2B. Indeed, impairment of autophagy and an increase in lysosomal degradative activity were found in cells expressing the mutant proteins. Recently, we described a novel <i>RAB7</i> mutation associated with predominantly motor CMT2 and impaired EGFR trafficking. With the aim to analyze the autophagy process and lysosomal activity in CMT2B fibroblasts carrying the p.K126R RAB7 novel mutation and to investigate further the causes of the different phenotype, we have performed Western blot, immunofluorescence and cytometric analyses monitoring autophagic markers and endocytic proteins. Moreover, we investigated lipophagy by analyzing accumulation of lipid droplets and their co-localization with endolysosomal degradative compartments. We found that cells expressing the RAB7<sup>K126R</sup> mutant protein were characterized by impairment of autophagy and lipophagy processes and by a moderate increase in lysosomal activity compared to the previously studied cells carrying the RAB7<sup>V162M</sup> mutation. Thus, we concluded that EGFR trafficking alterations and a moderate increase in lysosomal activity with concomitant impairment of autophagy could induce the specific predominantly motor phenotype observed in K126R patients. |
| first_indexed | 2024-03-10T00:03:12Z |
| format | Article |
| id | doaj.art-6376a66c70ea4191b806d103f7670bee |
| institution | Directory Open Access Journal |
| issn | 2073-4409 |
| language | English |
| last_indexed | 2024-03-10T00:03:12Z |
| publishDate | 2022-01-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cells |
| spelling | doaj.art-6376a66c70ea4191b806d103f7670bee2023-11-23T16:12:59ZengMDPI AGCells2073-44092022-01-0111349610.3390/cells11030496Autophagy and Lysosomal Functionality in CMT2B Fibroblasts Carrying the RAB7<sup>K126R</sup> MutationRoberta Romano0Victoria Stefania Del Fiore1Paola Saveri2Ilaria Elena Palamà3Chiara Pisciotta4Davide Pareyson5Cecilia Bucci6Flora Guerra7Department of Biological and Environmental Sciences and Technologies, University of Salento, Via Provinciale Lecce-Monteroni n. 165, 73100 Lecce, ItalyDepartment of Biological and Environmental Sciences and Technologies, University of Salento, Via Provinciale Lecce-Monteroni n. 165, 73100 Lecce, ItalyDepartment of Clinical Neurosciences, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, ItalyNanotechnology Institute, CNR-NANOTEC, Via Monteroni, 73100 Lecce, ItalyDepartment of Clinical Neurosciences, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, ItalyDepartment of Clinical Neurosciences, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, ItalyDepartment of Biological and Environmental Sciences and Technologies, University of Salento, Via Provinciale Lecce-Monteroni n. 165, 73100 Lecce, ItalyDepartment of Biological and Environmental Sciences and Technologies, University of Salento, Via Provinciale Lecce-Monteroni n. 165, 73100 Lecce, ItalyCharcot-Marie-Tooth type 2B (CMT2B) disease is a dominant axonal peripheral neuropathy caused by five mutations in the <i>RAB7A</i> gene. Autophagy and late endocytic trafficking were already characterized in CMT2B. Indeed, impairment of autophagy and an increase in lysosomal degradative activity were found in cells expressing the mutant proteins. Recently, we described a novel <i>RAB7</i> mutation associated with predominantly motor CMT2 and impaired EGFR trafficking. With the aim to analyze the autophagy process and lysosomal activity in CMT2B fibroblasts carrying the p.K126R RAB7 novel mutation and to investigate further the causes of the different phenotype, we have performed Western blot, immunofluorescence and cytometric analyses monitoring autophagic markers and endocytic proteins. Moreover, we investigated lipophagy by analyzing accumulation of lipid droplets and their co-localization with endolysosomal degradative compartments. We found that cells expressing the RAB7<sup>K126R</sup> mutant protein were characterized by impairment of autophagy and lipophagy processes and by a moderate increase in lysosomal activity compared to the previously studied cells carrying the RAB7<sup>V162M</sup> mutation. Thus, we concluded that EGFR trafficking alterations and a moderate increase in lysosomal activity with concomitant impairment of autophagy could induce the specific predominantly motor phenotype observed in K126R patients.https://www.mdpi.com/2073-4409/11/3/496Charcot-Marie-ToothRAB7Aautophagylysosomelipophagylipid droplets |
| spellingShingle | Roberta Romano Victoria Stefania Del Fiore Paola Saveri Ilaria Elena Palamà Chiara Pisciotta Davide Pareyson Cecilia Bucci Flora Guerra Autophagy and Lysosomal Functionality in CMT2B Fibroblasts Carrying the RAB7<sup>K126R</sup> Mutation Cells Charcot-Marie-Tooth RAB7A autophagy lysosome lipophagy lipid droplets |
| title | Autophagy and Lysosomal Functionality in CMT2B Fibroblasts Carrying the RAB7<sup>K126R</sup> Mutation |
| title_full | Autophagy and Lysosomal Functionality in CMT2B Fibroblasts Carrying the RAB7<sup>K126R</sup> Mutation |
| title_fullStr | Autophagy and Lysosomal Functionality in CMT2B Fibroblasts Carrying the RAB7<sup>K126R</sup> Mutation |
| title_full_unstemmed | Autophagy and Lysosomal Functionality in CMT2B Fibroblasts Carrying the RAB7<sup>K126R</sup> Mutation |
| title_short | Autophagy and Lysosomal Functionality in CMT2B Fibroblasts Carrying the RAB7<sup>K126R</sup> Mutation |
| title_sort | autophagy and lysosomal functionality in cmt2b fibroblasts carrying the rab7 sup k126r sup mutation |
| topic | Charcot-Marie-Tooth RAB7A autophagy lysosome lipophagy lipid droplets |
| url | https://www.mdpi.com/2073-4409/11/3/496 |
| work_keys_str_mv | AT robertaromano autophagyandlysosomalfunctionalityincmt2bfibroblastscarryingtherab7supk126rsupmutation AT victoriastefaniadelfiore autophagyandlysosomalfunctionalityincmt2bfibroblastscarryingtherab7supk126rsupmutation AT paolasaveri autophagyandlysosomalfunctionalityincmt2bfibroblastscarryingtherab7supk126rsupmutation AT ilariaelenapalama autophagyandlysosomalfunctionalityincmt2bfibroblastscarryingtherab7supk126rsupmutation AT chiarapisciotta autophagyandlysosomalfunctionalityincmt2bfibroblastscarryingtherab7supk126rsupmutation AT davidepareyson autophagyandlysosomalfunctionalityincmt2bfibroblastscarryingtherab7supk126rsupmutation AT ceciliabucci autophagyandlysosomalfunctionalityincmt2bfibroblastscarryingtherab7supk126rsupmutation AT floraguerra autophagyandlysosomalfunctionalityincmt2bfibroblastscarryingtherab7supk126rsupmutation |