Vps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis

Abstract Tuberous Sclerosis Complex (TSC), a rare genetic disorder with mechanistic target of rapamycin complex 1 (mTORC1) hyperactivation, is characterized by multi-organ hamartomatous benign tumors including brain, skin, kidney, and lung (Lymphangioleiomyomatosis). mTORC1 hyperactivation drives me...

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Autores principales:	Harilaos Filippakis, Amine Belaid, Brian Siroky, Constance Wu, Nicola Alesi, Thomas Hougard, Julie Nijmeh, Hilaire C. Lam, Elizabeth P. Henske
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Tuberous Sclerosis Complex TSC2-deficient Cells Dextran Uptake Mouse Embryonic Fibroblasts (MEFs) Ethylisopropylamiloride (EIPA) Medicine R Science Q
Acceso en línea:	https://doaj.org/article/65c5fa2a638943b98987048fb33e5747
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spelling	oai:doaj.org-article:65c5fa2a638943b98987048fb33e57472021-12-02T15:07:47ZVps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis10.1038/s41598-018-32256-x2045-2322https://doaj.org/article/65c5fa2a638943b98987048fb33e57472018-09-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-32256-xhttps://doaj.org/toc/2045-2322Abstract Tuberous Sclerosis Complex (TSC), a rare genetic disorder with mechanistic target of rapamycin complex 1 (mTORC1) hyperactivation, is characterized by multi-organ hamartomatous benign tumors including brain, skin, kidney, and lung (Lymphangioleiomyomatosis). mTORC1 hyperactivation drives metabolic reprogramming including glucose and glutamine utilization, protein, nucleic acid and lipid synthesis. To investigate the mechanisms of exogenous nutrients uptake in Tsc2-deficient cells, we measured dextran uptake, a polysaccharide internalized via macropinocytosis. Tsc2-deficient cells showed a striking increase in dextran uptake (3-fold, p < 0.0001) relative to Tsc2-expressing cells, which was decreased (3-fold, p < 0.0001) with mTOR inhibitor, Torin1. Pharmacologic and genetic inhibition of the lipid kinase Vps34 markedly abrogated uptake of Dextran in Tsc2-deficient cells. Macropinocytosis was further increased in Tsc2-deficient cells that lack autophagic mechanisms, suggesting that autophagy inhibition leads to dependence on exogenous nutrient uptake in Tsc2-deficient cells. Treatment with a macropinocytosis inhibitor, ethylisopropylamiloride (EIPA), resulted in selective growth inhibition of Atg5-deficient, Tsc2-deficient cells (50%, p < 0.0001). Genetic inhibition of autophagy (Atg5−/− MEFs) sensitized cells with Tsc2 downregulation to the Vps34 inhibitor, SAR405, resulting in growth inhibition (75%, p < 0.0001). Finally, genetic downregulation of Vps34 inhibited tumor growth and increased tumor latency in an in vivo xenograft model of TSC. Our findings show that macropinocytosis is upregulated with Tsc2-deficiency via a Vps34-dependent mechanism to support their anabolic state. The dependence of Tsc2-deficient cells on exogenous nutrients may provide novel approaches for the treatment of TSC.Harilaos FilippakisAmine BelaidBrian SirokyConstance WuNicola AlesiThomas HougardJulie NijmehHilaire C. LamElizabeth P. HenskeNature PortfolioarticleTuberous Sclerosis ComplexTSC2-deficient CellsDextran UptakeMouse Embryonic Fibroblasts (MEFs)Ethylisopropylamiloride (EIPA)MedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-10 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Tuberous Sclerosis Complex TSC2-deficient Cells Dextran Uptake Mouse Embryonic Fibroblasts (MEFs) Ethylisopropylamiloride (EIPA) Medicine R Science Q
spellingShingle	Tuberous Sclerosis Complex TSC2-deficient Cells Dextran Uptake Mouse Embryonic Fibroblasts (MEFs) Ethylisopropylamiloride (EIPA) Medicine R Science Q Harilaos Filippakis Amine Belaid Brian Siroky Constance Wu Nicola Alesi Thomas Hougard Julie Nijmeh Hilaire C. Lam Elizabeth P. Henske Vps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis
description	Abstract Tuberous Sclerosis Complex (TSC), a rare genetic disorder with mechanistic target of rapamycin complex 1 (mTORC1) hyperactivation, is characterized by multi-organ hamartomatous benign tumors including brain, skin, kidney, and lung (Lymphangioleiomyomatosis). mTORC1 hyperactivation drives metabolic reprogramming including glucose and glutamine utilization, protein, nucleic acid and lipid synthesis. To investigate the mechanisms of exogenous nutrients uptake in Tsc2-deficient cells, we measured dextran uptake, a polysaccharide internalized via macropinocytosis. Tsc2-deficient cells showed a striking increase in dextran uptake (3-fold, p < 0.0001) relative to Tsc2-expressing cells, which was decreased (3-fold, p < 0.0001) with mTOR inhibitor, Torin1. Pharmacologic and genetic inhibition of the lipid kinase Vps34 markedly abrogated uptake of Dextran in Tsc2-deficient cells. Macropinocytosis was further increased in Tsc2-deficient cells that lack autophagic mechanisms, suggesting that autophagy inhibition leads to dependence on exogenous nutrient uptake in Tsc2-deficient cells. Treatment with a macropinocytosis inhibitor, ethylisopropylamiloride (EIPA), resulted in selective growth inhibition of Atg5-deficient, Tsc2-deficient cells (50%, p < 0.0001). Genetic inhibition of autophagy (Atg5−/− MEFs) sensitized cells with Tsc2 downregulation to the Vps34 inhibitor, SAR405, resulting in growth inhibition (75%, p < 0.0001). Finally, genetic downregulation of Vps34 inhibited tumor growth and increased tumor latency in an in vivo xenograft model of TSC. Our findings show that macropinocytosis is upregulated with Tsc2-deficiency via a Vps34-dependent mechanism to support their anabolic state. The dependence of Tsc2-deficient cells on exogenous nutrients may provide novel approaches for the treatment of TSC.
format	article
author	Harilaos Filippakis Amine Belaid Brian Siroky Constance Wu Nicola Alesi Thomas Hougard Julie Nijmeh Hilaire C. Lam Elizabeth P. Henske
author_facet	Harilaos Filippakis Amine Belaid Brian Siroky Constance Wu Nicola Alesi Thomas Hougard Julie Nijmeh Hilaire C. Lam Elizabeth P. Henske
author_sort	Harilaos Filippakis
title	Vps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis
title_short	Vps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis
title_full	Vps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis
title_fullStr	Vps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis
title_full_unstemmed	Vps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis
title_sort	vps34-mediated macropinocytosis in tuberous sclerosis complex 2-deficient cells supports tumorigenesis
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/65c5fa2a638943b98987048fb33e5747
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Vps34-mediated macropinocytosis in Tuberous Sclerosis Complex 2-deficient cells supports tumorigenesis

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