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Perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition

Cbl (cobalamin) utilization as an enzyme cofactor is dependent on its efficient transit through lysosomes to the cytosol and mitochondria. We have previously proposed that pathophysiological perturbations in lysosomal function may inhibit intracellular Cbl transport with consequences for down-stream...

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Autores principales: Zhao, Hua, Ruberu, Kalani, Li, Hongyun, Garner, Brett
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Portland Press Ltd. 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3908614/
https://www.ncbi.nlm.nih.gov/pubmed/27919045
http://dx.doi.org/10.1042/BSR20130130
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author Zhao, Hua
Ruberu, Kalani
Li, Hongyun
Garner, Brett
author_facet Zhao, Hua
Ruberu, Kalani
Li, Hongyun
Garner, Brett
author_sort Zhao, Hua
collection PubMed
description Cbl (cobalamin) utilization as an enzyme cofactor is dependent on its efficient transit through lysosomes to the cytosol and mitochondria. We have previously proposed that pathophysiological perturbations in lysosomal function may inhibit intracellular Cbl transport with consequences for down-stream metabolic pathways. In the current study, we used both HT1080 fibroblasts and SH-SY5Y neurons to assess the impact that protease inhibitors, chloroquine and leupeptin (N-acetyl-L-leucyl-L-leucyl-L-argininal), have on the distribution of [(57)Co]Cbl in lysosomes, mitochondria and cytosol. Under standard cell culture conditions the distribution of [(57)Co]Cbl in both neurons and fibroblasts was ~5% in lysosomes, 14% in mitochondria and 81% in cytosol. Treatment of cells with either 25 μM chloroquine or 40 μM leupeptin for 48 h significantly increased the lysosomal [(57)Co]Cbl levels, by 4-fold in fibroblasts and 10-fold in neurons, and this was associated with reduced cytosolic and mitochondrial [(57)Co]Cbl concentrations. Based on Western blotting of LAMP2 in fractions recovered from an OptiPrep density gradient, lysosomal Cbl trapping was associated with an expansion of the lysosomal compartment and an increase in a subpopulation of lysosomes with increased size and density. Moreover, the decreased mitochondrial Cbl that was associated with lysosomal Cbl trapping was correlated with decreased incorporation of [(14)C] propionate into cellular proteins/macromolecules, indicating an inhibition of Cbl-dependent Mm-CoA (methylmalonyl-coenzyme A) mutase activity. These results add support to the idea that lysosomal dysfunction may significantly impact upon Cbl transport and utilization.
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spelling pubmed-39086142014-02-03 Perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition Zhao, Hua Ruberu, Kalani Li, Hongyun Garner, Brett Biosci Rep Original Paper Cbl (cobalamin) utilization as an enzyme cofactor is dependent on its efficient transit through lysosomes to the cytosol and mitochondria. We have previously proposed that pathophysiological perturbations in lysosomal function may inhibit intracellular Cbl transport with consequences for down-stream metabolic pathways. In the current study, we used both HT1080 fibroblasts and SH-SY5Y neurons to assess the impact that protease inhibitors, chloroquine and leupeptin (N-acetyl-L-leucyl-L-leucyl-L-argininal), have on the distribution of [(57)Co]Cbl in lysosomes, mitochondria and cytosol. Under standard cell culture conditions the distribution of [(57)Co]Cbl in both neurons and fibroblasts was ~5% in lysosomes, 14% in mitochondria and 81% in cytosol. Treatment of cells with either 25 μM chloroquine or 40 μM leupeptin for 48 h significantly increased the lysosomal [(57)Co]Cbl levels, by 4-fold in fibroblasts and 10-fold in neurons, and this was associated with reduced cytosolic and mitochondrial [(57)Co]Cbl concentrations. Based on Western blotting of LAMP2 in fractions recovered from an OptiPrep density gradient, lysosomal Cbl trapping was associated with an expansion of the lysosomal compartment and an increase in a subpopulation of lysosomes with increased size and density. Moreover, the decreased mitochondrial Cbl that was associated with lysosomal Cbl trapping was correlated with decreased incorporation of [(14)C] propionate into cellular proteins/macromolecules, indicating an inhibition of Cbl-dependent Mm-CoA (methylmalonyl-coenzyme A) mutase activity. These results add support to the idea that lysosomal dysfunction may significantly impact upon Cbl transport and utilization. Portland Press Ltd. 2014-01-31 /pmc/articles/PMC3908614/ /pubmed/27919045 http://dx.doi.org/10.1042/BSR20130130 Text en © 2014 The author(s) has paid for this article to be freely available under the terms of the Creative Commons Attribution Licence (CC-BY)(http://creativecommons.org/licenses/by/3.0/) which permits unrestricted use, distribution and reproduction in any medium, provided the original work is properly cited. http://creativecommons.org/licenses/by/3.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution Licence (CC-BY) (http://creativecommons.org/licenses/by/3.0/) which permits unrestricted use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Original Paper
Zhao, Hua
Ruberu, Kalani
Li, Hongyun
Garner, Brett
Perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition
title Perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition
title_full Perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition
title_fullStr Perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition
title_full_unstemmed Perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition
title_short Perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition
title_sort perturbation of neuronal cobalamin transport by lysosomal enzyme inhibition
topic Original Paper
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3908614/
https://www.ncbi.nlm.nih.gov/pubmed/27919045
http://dx.doi.org/10.1042/BSR20130130
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