In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation
Hypoxia-inducible factor (HIF) mediates a broad, conserved adaptive response to hypoxia, and the HIF pathway is a potential therapeutic target in cerebral ischemia. This study investigated the mechanism by which in vitro ischemia (oxygen-glucose deprivation; OGD) affects canonical hypoxic HIF-1α sta...
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2013
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| Online Access: | http://hdl.handle.net/10725/6137 http://dx.doi.org/10.1002/jnr.23204 http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.php |
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| _version_ | 1864513466150158336 |
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| author | Sleiman, Sama F. |
| author_facet | Sleiman, Sama F. |
| author_role | author |
| dc.creator.none.fl_str_mv | Sleiman, Sama F. |
| dc.date.none.fl_str_mv | 2013 2017-09-05T13:31:35Z 2017-09-05T13:31:35Z 2017-09-05 |
| dc.identifier.none.fl_str_mv | 1097-4547 http://hdl.handle.net/10725/6137 http://dx.doi.org/10.1002/jnr.23204 Karuppagounder, S. S., Basso, M., Sleiman, S. F., Ma, T. C., Speer, R. E., Smirnova, N. A., ... & Ratan, R. R. (2013). In vitro ischemia suppresses hypoxic induction of hypoxia‐inducible factor‐1α by inhibition of synthesis and not enhanced degradation. Journal of neuroscience research, 91(8), 1066-1075. http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.php file:///C:/Users/rola.habre/Downloads/Karuppagounder_et_al-2013-Journal_of_Neuroscience_Research.pdf |
| dc.language.none.fl_str_mv | en |
| dc.relation.none.fl_str_mv | Journal of Neuroscience Research |
| dc.rights.*.fl_str_mv | info:eu-repo/semantics/openAccess |
| dc.title.none.fl_str_mv | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation |
| dc.type.none.fl_str_mv | Article info:eu-repo/semantics/publishedVersion info:eu-repo/semantics/article |
| description | Hypoxia-inducible factor (HIF) mediates a broad, conserved adaptive response to hypoxia, and the HIF pathway is a potential therapeutic target in cerebral ischemia. This study investigated the mechanism by which in vitro ischemia (oxygen-glucose deprivation; OGD) affects canonical hypoxic HIF-1α stabilization. We validated the use of a reporter containing the oxygen-dependent degradation domain of HIF-1α fused to firefly luciferase (ODD-luc) to monitor quantitatively distinct biochemical events leading to hypoxic HIF-1α expression or stabilization in a human neuroblastoma cell line (SH-SY5Y). When OGD was imposed following a 2-hr hypoxic stabilization of ODD-luc, the levels of the reporter were reduced, consistent with prior models proposing that OGD enhances HIF prolylhydroxylase (PHD) activity. Surprisingly, PHD inhibitors and proteasome inhibitors do not stabilize ODD-luc in OGD. Furthermore, OGD does not affect the half-life of ODD-luc protein following hypoxia, suggesting that OGD abrogates hypoxic HIF-1α induction by reducing HIF-1α synthesis rather than by enhancing its degradation. We observed ATP depletion under OGD vs. hypoxia and propose that ATP depletion enhances translational suppression, overcoming the selective synthesis of HIF concurrent with global decreases in protein synthesis in hypoxia. Taken together, these findings biochemically characterize a practical reporter for monitoring HIF-1α levels and support a novel model for HIF regulation in an in vitro model of human ischemia. |
| eu_rights_str_mv | openAccess |
| format | article |
| id | LAURepo_f4044ebedfdeed6f9734c67ba23c5016 |
| identifier_str_mv | 1097-4547 Karuppagounder, S. S., Basso, M., Sleiman, S. F., Ma, T. C., Speer, R. E., Smirnova, N. A., ... & Ratan, R. R. (2013). In vitro ischemia suppresses hypoxic induction of hypoxia‐inducible factor‐1α by inhibition of synthesis and not enhanced degradation. Journal of neuroscience research, 91(8), 1066-1075. file:///C:/Users/rola.habre/Downloads/Karuppagounder_et_al-2013-Journal_of_Neuroscience_Research.pdf |
| language_invalid_str_mv | en |
| network_acronym_str | LAURepo |
| network_name_str | Lebanese American University repository |
| oai_identifier_str | oai:laur.lau.edu.lb:10725/6137 |
| publishDate | 2013 |
| repository.mail.fl_str_mv | |
| repository.name.fl_str_mv | |
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| spelling | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradationSleiman, Sama F.Hypoxia-inducible factor (HIF) mediates a broad, conserved adaptive response to hypoxia, and the HIF pathway is a potential therapeutic target in cerebral ischemia. This study investigated the mechanism by which in vitro ischemia (oxygen-glucose deprivation; OGD) affects canonical hypoxic HIF-1α stabilization. We validated the use of a reporter containing the oxygen-dependent degradation domain of HIF-1α fused to firefly luciferase (ODD-luc) to monitor quantitatively distinct biochemical events leading to hypoxic HIF-1α expression or stabilization in a human neuroblastoma cell line (SH-SY5Y). When OGD was imposed following a 2-hr hypoxic stabilization of ODD-luc, the levels of the reporter were reduced, consistent with prior models proposing that OGD enhances HIF prolylhydroxylase (PHD) activity. Surprisingly, PHD inhibitors and proteasome inhibitors do not stabilize ODD-luc in OGD. Furthermore, OGD does not affect the half-life of ODD-luc protein following hypoxia, suggesting that OGD abrogates hypoxic HIF-1α induction by reducing HIF-1α synthesis rather than by enhancing its degradation. We observed ATP depletion under OGD vs. hypoxia and propose that ATP depletion enhances translational suppression, overcoming the selective synthesis of HIF concurrent with global decreases in protein synthesis in hypoxia. Taken together, these findings biochemically characterize a practical reporter for monitoring HIF-1α levels and support a novel model for HIF regulation in an in vitro model of human ischemia.PublishedN/A2017-09-05T13:31:35Z2017-09-05T13:31:35Z20132017-09-05Articleinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/article1097-4547http://hdl.handle.net/10725/6137http://dx.doi.org/10.1002/jnr.23204Karuppagounder, S. S., Basso, M., Sleiman, S. F., Ma, T. C., Speer, R. E., Smirnova, N. A., ... & Ratan, R. R. (2013). In vitro ischemia suppresses hypoxic induction of hypoxia‐inducible factor‐1α by inhibition of synthesis and not enhanced degradation. Journal of neuroscience research, 91(8), 1066-1075.http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.phpfile:///C:/Users/rola.habre/Downloads/Karuppagounder_et_al-2013-Journal_of_Neuroscience_Research.pdfenJournal of Neuroscience Researchinfo:eu-repo/semantics/openAccessoai:laur.lau.edu.lb:10725/61372021-03-19T10:00:52Z |
| spellingShingle | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation Sleiman, Sama F. |
| status_str | publishedVersion |
| title | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation |
| title_full | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation |
| title_fullStr | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation |
| title_full_unstemmed | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation |
| title_short | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation |
| title_sort | In vitro ischemia suppresses hypoxic induction of hypoxia-inducible factor-1α by inhibition of synthesis and not enhanced degradation |
| url | http://hdl.handle.net/10725/6137 http://dx.doi.org/10.1002/jnr.23204 http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.php |