PLEKHG5 regulates autophagy, survival and MGMT expression in U251-MG glioblastoma cells
A signalling pathway involving PLEKHG5 (guanine exchange factor) for the Ras superfamily member RAB26 to transcription factor NF-κB was discovered in autophagy. PLEKHG5 was reported in glioblastoma multiforme (GBM) and correlates with patient survival. Thus, the generation of a cellular model for understanding PLEKHG5 signalling is the study purpose. We generated a CRISPR/Cas9-mediated knockout of PLEKHG5 in U251-MG glioblastoma cells and analysed resulting changes. Next, we used a mRFP-GFP-LC3+ reporter for visualisation of autophagic defects and rescued the phenotype of PLEKHG5 wildtype via transduction of a constitutively active RAB26QL-plasmid. Effects of overexpressing RAB26 were investigated and correlated with the O6-methylguanine-DNA methyltransferase (MGMT) and cellular survival. PLEKHG5 knockout showed changes in morphology, loss of filopodia and higher population doubling times. Accumulation of autolysosomes was resulted by decreased LAMP-1 in PLEKHG5-deficient cells. Rescue of PLEKHG5-/- restored the downregulation of RhoA activity, showed faster response to tumour necrosis factor and better cellular fitness. MGMT expression was activated after RAB26 overexpression compared to non-transduced cells. Survival of PLEKHG5 knockout was rescued together with sensitivity to temozolomide by RAB26QL. This study provides new insights in the PLEKHG5/RAB26 signalling within U251-MG cells, which suggests potential therapeutic strategies in other glioma cells and further in primary GBM.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2020 |
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| Erschienen: |
2020 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:10 |
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| Enthalten in: |
Scientific reports - 10(2020), 1 vom: 14. Dez., Seite 21858 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Witte, Kaya Elisa [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 28.04.2021 Date Revised 10.11.2023 published: Electronic Citation Status MEDLINE |
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| doi: |
10.1038/s41598-020-77958-3 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM31883703X |
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| 100 | 1 | |a Witte, Kaya Elisa |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a PLEKHG5 regulates autophagy, survival and MGMT expression in U251-MG glioblastoma cells |
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| 500 | |a Date Completed 28.04.2021 | ||
| 500 | |a Date Revised 10.11.2023 | ||
| 500 | |a published: Electronic | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a A signalling pathway involving PLEKHG5 (guanine exchange factor) for the Ras superfamily member RAB26 to transcription factor NF-κB was discovered in autophagy. PLEKHG5 was reported in glioblastoma multiforme (GBM) and correlates with patient survival. Thus, the generation of a cellular model for understanding PLEKHG5 signalling is the study purpose. We generated a CRISPR/Cas9-mediated knockout of PLEKHG5 in U251-MG glioblastoma cells and analysed resulting changes. Next, we used a mRFP-GFP-LC3+ reporter for visualisation of autophagic defects and rescued the phenotype of PLEKHG5 wildtype via transduction of a constitutively active RAB26QL-plasmid. Effects of overexpressing RAB26 were investigated and correlated with the O6-methylguanine-DNA methyltransferase (MGMT) and cellular survival. PLEKHG5 knockout showed changes in morphology, loss of filopodia and higher population doubling times. Accumulation of autolysosomes was resulted by decreased LAMP-1 in PLEKHG5-deficient cells. Rescue of PLEKHG5-/- restored the downregulation of RhoA activity, showed faster response to tumour necrosis factor and better cellular fitness. MGMT expression was activated after RAB26 overexpression compared to non-transduced cells. Survival of PLEKHG5 knockout was rescued together with sensitivity to temozolomide by RAB26QL. This study provides new insights in the PLEKHG5/RAB26 signalling within U251-MG cells, which suggests potential therapeutic strategies in other glioma cells and further in primary GBM | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 7 | |a Guanine Nucleotide Exchange Factors |2 NLM | |
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| 700 | 1 | |a Slotta, Carsten |e verfasserin |4 aut | |
| 700 | 1 | |a Lütkemeyer, Melanie |e verfasserin |4 aut | |
| 700 | 1 | |a Kitke, Angelika |e verfasserin |4 aut | |
| 700 | 1 | |a Coras, Roland |e verfasserin |4 aut | |
| 700 | 1 | |a Simon, Matthias |e verfasserin |4 aut | |
| 700 | 1 | |a Kaltschmidt, Christian |e verfasserin |4 aut | |
| 700 | 1 | |a Kaltschmidt, Barbara |e verfasserin |4 aut | |
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