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RNA Interference Screening Identifies Novel Roles for RhoBTB1 and RhoBTB3 in Membrane Trafficking Events in Mammalian Cells

In the endomembrane system of mammalian cells, membrane traffic processes require a high degree of regulation in order to ensure their specificity. The range of molecules that participate in trafficking events is truly vast, and much attention to date has been given to the Rab family of small GTPase...

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Autores principales: Long, Maeve, Kranjc, Tilen, Mysior, Margaritha M., Simpson, Jeremy C.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7291084/
https://www.ncbi.nlm.nih.gov/pubmed/32354068
http://dx.doi.org/10.3390/cells9051089
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author Long, Maeve
Kranjc, Tilen
Mysior, Margaritha M.
Simpson, Jeremy C.
author_facet Long, Maeve
Kranjc, Tilen
Mysior, Margaritha M.
Simpson, Jeremy C.
author_sort Long, Maeve
collection PubMed
description In the endomembrane system of mammalian cells, membrane traffic processes require a high degree of regulation in order to ensure their specificity. The range of molecules that participate in trafficking events is truly vast, and much attention to date has been given to the Rab family of small GTPases. However, in recent years, a role in membrane traffic for members of the Rho GTPase family, in particular Cdc42, has emerged. This prompted us to develop and apply an image-based high-content screen, initially focussing on the Golgi complex, using RNA interference to systematically perturb each of the 21 Rho family members and assess their importance to the overall organisation of this organelle. Analysis of our data revealed previously unreported roles for two atypical Rho family members, RhoBTB1 and RhoBTB3, in membrane traffic events. We find that depletion of RhoBTB3 affects the morphology of the Golgi complex and causes changes in the trafficking speeds of carriers operating at the interface of the Golgi and endoplasmic reticulum. In addition, RhoBTB3 was found to be present on these carriers. Depletion of RhoBTB1 was also found to cause a disturbance to the Golgi architecture, however, this phenotype seems to be linked to endocytosis and retrograde traffic pathways. RhoBTB1 was found to be associated with early endosomal intermediates, and changes in the levels of RhoBTB1 not only caused profound changes to the organisation and distribution of endosomes and lysosomes, but also resulted in defects in the delivery of two different classes of cargo molecules to downstream compartments. Together, our data reveal new roles for these atypical Rho family members in the endomembrane system.
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spelling pubmed-72910842020-06-17 RNA Interference Screening Identifies Novel Roles for RhoBTB1 and RhoBTB3 in Membrane Trafficking Events in Mammalian Cells Long, Maeve Kranjc, Tilen Mysior, Margaritha M. Simpson, Jeremy C. Cells Article In the endomembrane system of mammalian cells, membrane traffic processes require a high degree of regulation in order to ensure their specificity. The range of molecules that participate in trafficking events is truly vast, and much attention to date has been given to the Rab family of small GTPases. However, in recent years, a role in membrane traffic for members of the Rho GTPase family, in particular Cdc42, has emerged. This prompted us to develop and apply an image-based high-content screen, initially focussing on the Golgi complex, using RNA interference to systematically perturb each of the 21 Rho family members and assess their importance to the overall organisation of this organelle. Analysis of our data revealed previously unreported roles for two atypical Rho family members, RhoBTB1 and RhoBTB3, in membrane traffic events. We find that depletion of RhoBTB3 affects the morphology of the Golgi complex and causes changes in the trafficking speeds of carriers operating at the interface of the Golgi and endoplasmic reticulum. In addition, RhoBTB3 was found to be present on these carriers. Depletion of RhoBTB1 was also found to cause a disturbance to the Golgi architecture, however, this phenotype seems to be linked to endocytosis and retrograde traffic pathways. RhoBTB1 was found to be associated with early endosomal intermediates, and changes in the levels of RhoBTB1 not only caused profound changes to the organisation and distribution of endosomes and lysosomes, but also resulted in defects in the delivery of two different classes of cargo molecules to downstream compartments. Together, our data reveal new roles for these atypical Rho family members in the endomembrane system. MDPI 2020-04-28 /pmc/articles/PMC7291084/ /pubmed/32354068 http://dx.doi.org/10.3390/cells9051089 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Long, Maeve
Kranjc, Tilen
Mysior, Margaritha M.
Simpson, Jeremy C.
RNA Interference Screening Identifies Novel Roles for RhoBTB1 and RhoBTB3 in Membrane Trafficking Events in Mammalian Cells
title RNA Interference Screening Identifies Novel Roles for RhoBTB1 and RhoBTB3 in Membrane Trafficking Events in Mammalian Cells
title_full RNA Interference Screening Identifies Novel Roles for RhoBTB1 and RhoBTB3 in Membrane Trafficking Events in Mammalian Cells
title_fullStr RNA Interference Screening Identifies Novel Roles for RhoBTB1 and RhoBTB3 in Membrane Trafficking Events in Mammalian Cells
title_full_unstemmed RNA Interference Screening Identifies Novel Roles for RhoBTB1 and RhoBTB3 in Membrane Trafficking Events in Mammalian Cells
title_short RNA Interference Screening Identifies Novel Roles for RhoBTB1 and RhoBTB3 in Membrane Trafficking Events in Mammalian Cells
title_sort rna interference screening identifies novel roles for rhobtb1 and rhobtb3 in membrane trafficking events in mammalian cells
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7291084/
https://www.ncbi.nlm.nih.gov/pubmed/32354068
http://dx.doi.org/10.3390/cells9051089
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