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Calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma

Glioblastoma (GBM) is an aggressive, malignant brain tumor that inevitably develops resistance to conventional chemotherapy and radiation treatments. In order to identify signaling pathways involved in the development of radiation resistance, we performed mass spectrometry-based phospho-proteomic pr...

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Autores principales: Bassett, Emily A., Palanichamy, Kamalakannan, Pearson, Mitchell, McElroy, Joseph P., Haque, Saikh Jaharul, Bell, Erica Hlavin, Chakravarti, Arnab
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Impact Journals LLC 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5865692/
https://www.ncbi.nlm.nih.gov/pubmed/29581866
http://dx.doi.org/10.18632/oncotarget.24523
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author Bassett, Emily A.
Palanichamy, Kamalakannan
Pearson, Mitchell
McElroy, Joseph P.
Haque, Saikh Jaharul
Bell, Erica Hlavin
Chakravarti, Arnab
author_facet Bassett, Emily A.
Palanichamy, Kamalakannan
Pearson, Mitchell
McElroy, Joseph P.
Haque, Saikh Jaharul
Bell, Erica Hlavin
Chakravarti, Arnab
author_sort Bassett, Emily A.
collection PubMed
description Glioblastoma (GBM) is an aggressive, malignant brain tumor that inevitably develops resistance to conventional chemotherapy and radiation treatments. In order to identify signaling pathways involved in the development of radiation resistance, we performed mass spectrometry-based phospho-proteomic profiling of GBM cell lines and normal human astrocytes before and after radiation treatment. We found radiation induced phosphorylation of a number of proteins including calpastatin, specifically in GBM stem cells (GSCs). Herein, we focused on calpastatin, an endogenous inhibitor of calpain proteases. Radiation-induced phosphorylation of calpastatin at Ser-633 within the inhibitory domain was validated with a phospho-specific antibody. In order to test the functional significance of phosphorylated calpastatin, we utilized site-directed mutagenesis to generate phospho-inactive (Ser633Ala) and phospho-mimetic (Ser633Glu) mutant calpastatin. GBM cell lines stably expressing the mutant calpastatin showed that phosphorylation was necessary for radiation-induced calpain activation. We also showed that casein kinase 2, a pro-survival kinase overexpressed in many cancer types, phosphorylated calpastatin at Ser-633. Our results indicate that calpastatin phosphorylation promotes radiation resistance in GBM cells by increasing the activity of calpain proteases, which are known to promote survival and invasion in cancer.
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spelling pubmed-58656922018-03-26 Calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma Bassett, Emily A. Palanichamy, Kamalakannan Pearson, Mitchell McElroy, Joseph P. Haque, Saikh Jaharul Bell, Erica Hlavin Chakravarti, Arnab Oncotarget Research Paper Glioblastoma (GBM) is an aggressive, malignant brain tumor that inevitably develops resistance to conventional chemotherapy and radiation treatments. In order to identify signaling pathways involved in the development of radiation resistance, we performed mass spectrometry-based phospho-proteomic profiling of GBM cell lines and normal human astrocytes before and after radiation treatment. We found radiation induced phosphorylation of a number of proteins including calpastatin, specifically in GBM stem cells (GSCs). Herein, we focused on calpastatin, an endogenous inhibitor of calpain proteases. Radiation-induced phosphorylation of calpastatin at Ser-633 within the inhibitory domain was validated with a phospho-specific antibody. In order to test the functional significance of phosphorylated calpastatin, we utilized site-directed mutagenesis to generate phospho-inactive (Ser633Ala) and phospho-mimetic (Ser633Glu) mutant calpastatin. GBM cell lines stably expressing the mutant calpastatin showed that phosphorylation was necessary for radiation-induced calpain activation. We also showed that casein kinase 2, a pro-survival kinase overexpressed in many cancer types, phosphorylated calpastatin at Ser-633. Our results indicate that calpastatin phosphorylation promotes radiation resistance in GBM cells by increasing the activity of calpain proteases, which are known to promote survival and invasion in cancer. Impact Journals LLC 2018-02-19 /pmc/articles/PMC5865692/ /pubmed/29581866 http://dx.doi.org/10.18632/oncotarget.24523 Text en Copyright: © 2018 Bassett et al. http://creativecommons.org/licenses/by/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License 3.0 (http://creativecommons.org/licenses/by/3.0/) (CC BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Paper
Bassett, Emily A.
Palanichamy, Kamalakannan
Pearson, Mitchell
McElroy, Joseph P.
Haque, Saikh Jaharul
Bell, Erica Hlavin
Chakravarti, Arnab
Calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma
title Calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma
title_full Calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma
title_fullStr Calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma
title_full_unstemmed Calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma
title_short Calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma
title_sort calpastatin phosphorylation regulates radiation-induced calpain activity in glioblastoma
topic Research Paper
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5865692/
https://www.ncbi.nlm.nih.gov/pubmed/29581866
http://dx.doi.org/10.18632/oncotarget.24523
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