Calcineurin inhibitor-induced and Ras-mediated overexpression of VEGF in renal cancer cells involves mTOR through the regulation of PRAS40.

Malignancy is a major problem in patients treated with immunosuppressive agents. We have demonstrated that treatment with calcineurin inhibitors (CNIs) can induce the activation of proto-oncogenic Ras, and may promote a rapid progression of human renal cancer through the overexpression of vascular e...

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Main Authors: Aninda Basu, Pallavi Banerjee, Alan G Contreras, Evelyn Flynn, Soumitro Pal
Format: Article
Language:English
Published: Public Library of Science (PLoS) 2011-01-01
Series:PLoS ONE
Online Access:http://europepmc.org/articles/PMC3160347?pdf=render
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spelling doaj-56c2393df95b442db39fdd9ce2b1f6bb2020-11-25T01:38:17ZengPublic Library of Science (PLoS)PLoS ONE1932-62032011-01-0168e2391910.1371/journal.pone.0023919Calcineurin inhibitor-induced and Ras-mediated overexpression of VEGF in renal cancer cells involves mTOR through the regulation of PRAS40.Aninda BasuPallavi BanerjeeAlan G ContrerasEvelyn FlynnSoumitro PalMalignancy is a major problem in patients treated with immunosuppressive agents. We have demonstrated that treatment with calcineurin inhibitors (CNIs) can induce the activation of proto-oncogenic Ras, and may promote a rapid progression of human renal cancer through the overexpression of vascular endothelial growth factor (VEGF). Interestingly, we found that CNI-induced VEGF overexpression and cancer cell proliferation was inhibited by rapamycin treatment, indicating potential involvement of the mammalian target of rapamycin (mTOR) pathway in this tumorigenic process. Here, we examined the role of mTOR pathway in mediating CNI- and Ras-induced overexpression of VEGF in human renal cancer cells (786-0 and Caki-1). We found that the knockdown of raptor (using siRNA) significantly decreased CNI-induced VEGF promoter activity as observed by promoter-luciferase assay, suggesting the role of mTOR complex1 (mTORC1) in CNI-induced VEGF transcription. It is known that mTOR becomes activated following phosphorylation of its negative regulator PRAS40, which is a part of mTORC1. We observed that CNI treatment and activation of H-Ras (through transfection of an active H-Ras plasmid) markedly increased the phosphorylation of PRAS40, and the transfection of cells using a dominant-negative plasmid of Ras, significantly decreased PRAS40 phosphorylation. Protein kinase C (PKC)-ζ and PKC-δ, which are critical intermediary signaling molecules for CNI-induced tumorigenic pathway, formed complex with PRAS40; and we found that the CNI treatment increased the complex formation between PRAS40 and PKC, particularly (PKC)-ζ. Inhibition of PKC activity using pharmacological inhibitor markedly decreased H-Ras-induced phosphorylation of PRAS40. The overexpression of PRAS40 in renal cancer cells significantly down-regulated CNI- and H-Ras-induced VEGF transcriptional activation. Finally, it was observed that CNI treatment increased the expression of phosho-PRAS40 in renal tumor tissues in vivo. Together, the phosphorylation of PRAS40 is critical for the activation of mTOR in CNI-induced VEGF overexpression and renal cancer progression.http://europepmc.org/articles/PMC3160347?pdf=render
collection DOAJ
language English
format Article
sources DOAJ
author Aninda Basu
Pallavi Banerjee
Alan G Contreras
Evelyn Flynn
Soumitro Pal
spellingShingle Aninda Basu
Pallavi Banerjee
Alan G Contreras
Evelyn Flynn
Soumitro Pal
Calcineurin inhibitor-induced and Ras-mediated overexpression of VEGF in renal cancer cells involves mTOR through the regulation of PRAS40.
PLoS ONE
author_facet Aninda Basu
Pallavi Banerjee
Alan G Contreras
Evelyn Flynn
Soumitro Pal
author_sort Aninda Basu
title Calcineurin inhibitor-induced and Ras-mediated overexpression of VEGF in renal cancer cells involves mTOR through the regulation of PRAS40.
title_short Calcineurin inhibitor-induced and Ras-mediated overexpression of VEGF in renal cancer cells involves mTOR through the regulation of PRAS40.
title_full Calcineurin inhibitor-induced and Ras-mediated overexpression of VEGF in renal cancer cells involves mTOR through the regulation of PRAS40.
title_fullStr Calcineurin inhibitor-induced and Ras-mediated overexpression of VEGF in renal cancer cells involves mTOR through the regulation of PRAS40.
title_full_unstemmed Calcineurin inhibitor-induced and Ras-mediated overexpression of VEGF in renal cancer cells involves mTOR through the regulation of PRAS40.
title_sort calcineurin inhibitor-induced and ras-mediated overexpression of vegf in renal cancer cells involves mtor through the regulation of pras40.
publisher Public Library of Science (PLoS)
series PLoS ONE
issn 1932-6203
publishDate 2011-01-01
description Malignancy is a major problem in patients treated with immunosuppressive agents. We have demonstrated that treatment with calcineurin inhibitors (CNIs) can induce the activation of proto-oncogenic Ras, and may promote a rapid progression of human renal cancer through the overexpression of vascular endothelial growth factor (VEGF). Interestingly, we found that CNI-induced VEGF overexpression and cancer cell proliferation was inhibited by rapamycin treatment, indicating potential involvement of the mammalian target of rapamycin (mTOR) pathway in this tumorigenic process. Here, we examined the role of mTOR pathway in mediating CNI- and Ras-induced overexpression of VEGF in human renal cancer cells (786-0 and Caki-1). We found that the knockdown of raptor (using siRNA) significantly decreased CNI-induced VEGF promoter activity as observed by promoter-luciferase assay, suggesting the role of mTOR complex1 (mTORC1) in CNI-induced VEGF transcription. It is known that mTOR becomes activated following phosphorylation of its negative regulator PRAS40, which is a part of mTORC1. We observed that CNI treatment and activation of H-Ras (through transfection of an active H-Ras plasmid) markedly increased the phosphorylation of PRAS40, and the transfection of cells using a dominant-negative plasmid of Ras, significantly decreased PRAS40 phosphorylation. Protein kinase C (PKC)-ζ and PKC-δ, which are critical intermediary signaling molecules for CNI-induced tumorigenic pathway, formed complex with PRAS40; and we found that the CNI treatment increased the complex formation between PRAS40 and PKC, particularly (PKC)-ζ. Inhibition of PKC activity using pharmacological inhibitor markedly decreased H-Ras-induced phosphorylation of PRAS40. The overexpression of PRAS40 in renal cancer cells significantly down-regulated CNI- and H-Ras-induced VEGF transcriptional activation. Finally, it was observed that CNI treatment increased the expression of phosho-PRAS40 in renal tumor tissues in vivo. Together, the phosphorylation of PRAS40 is critical for the activation of mTOR in CNI-induced VEGF overexpression and renal cancer progression.
url http://europepmc.org/articles/PMC3160347?pdf=render
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