EPAS1 trans-activation during hypoxia requires p42/p44 MAPK

P. William Conrad, Thomas L. Freeman, Dana Beitner-Johnson, David E. Millhorn

Research output: Contribution to journalArticle

142 Citations (Scopus)

Abstract

Hypoxia is a common environmental stress that regulates gene expression and cell function. A number of hypoxia-regulated transcription factors have been identified and have been shown to play critical roles in mediating cellular responses to hypoxia. One of these is the endothelial PAS-domain protein 1 (EPAS1/HIF2-α/HLF/HRF). This protein is 48% homologous to hypoxia- inducible factor 1-α (HIF1-α). To date, virtually nothing is known about the signaling pathways that lead to either EPAS1 or HIF1-α activation. Here we show that EPAS1 is phosphorylated when PC12 cells are exposed to hypoxia and that p42/p44 MAPK is a critical mediator of EPAS1 activation. Pretreatment of PC12 cells with the MEK inhibitor, PD98059, completely blocked hypoxia-induced trans-activation of a hypoxia response element (HRE) reporter gene by transfected EPAS1. Likewise, expression of a constitutively active MEK1 mimicked the effects of hypoxia on HRE reporter gene expression. However, pretreatment with PD98059 had no effect on EPAS1 phosphorylation during hypoxia, suggesting that MAPK targets other proteins that are critical for the trans-activation of EPAS1. We further show that hypoxia-induced trans-activation of EPAS1 is independent of Ras. Finally, pretreatment with calmodulin antagonists nearly completely blocked both the hypoxia-induced phosphorylation of MAPK and the EPAS1 trans-activation of HRE-Luc. These results demonstrate that the MAPK pathway is a critical mediator of EPAS1 activation and that activation of MAPK and EPAS1 occurs through a calmodulin- sensitive pathway and not through the GTPase, Ras. These results are the first to identify a specific signaling pathway involved in EPAS1 activation.

Original languageEnglish (US)
Pages (from-to)33709-33713
Number of pages5
JournalJournal of Biological Chemistry
Volume274
Issue number47
DOIs
StatePublished - Nov 19 1999

Fingerprint

Mitogen-Activated Protein Kinase 3
Mitogen-Activated Protein Kinase 1
Chemical activation
Response Elements
Hypoxia-Inducible Factor 1
Phosphorylation
Calmodulin
Gene expression
PC12 Cells
Reporter Genes
Hypoxia
Proteins
GTP Phosphohydrolases
Mitogen-Activated Protein Kinase Kinases
Gene Expression
Transcription Factors
Genes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

EPAS1 trans-activation during hypoxia requires p42/p44 MAPK. / Conrad, P. William; Freeman, Thomas L.; Beitner-Johnson, Dana; Millhorn, David E.

In: Journal of Biological Chemistry, Vol. 274, No. 47, 19.11.1999, p. 33709-33713.

Research output: Contribution to journalArticle

Conrad, P. William ; Freeman, Thomas L. ; Beitner-Johnson, Dana ; Millhorn, David E. / EPAS1 trans-activation during hypoxia requires p42/p44 MAPK. In: Journal of Biological Chemistry. 1999 ; Vol. 274, No. 47. pp. 33709-33713.
@article{d5aa7ed1048d4613bbf041556017d744,
title = "EPAS1 trans-activation during hypoxia requires p42/p44 MAPK",
abstract = "Hypoxia is a common environmental stress that regulates gene expression and cell function. A number of hypoxia-regulated transcription factors have been identified and have been shown to play critical roles in mediating cellular responses to hypoxia. One of these is the endothelial PAS-domain protein 1 (EPAS1/HIF2-α/HLF/HRF). This protein is 48{\%} homologous to hypoxia- inducible factor 1-α (HIF1-α). To date, virtually nothing is known about the signaling pathways that lead to either EPAS1 or HIF1-α activation. Here we show that EPAS1 is phosphorylated when PC12 cells are exposed to hypoxia and that p42/p44 MAPK is a critical mediator of EPAS1 activation. Pretreatment of PC12 cells with the MEK inhibitor, PD98059, completely blocked hypoxia-induced trans-activation of a hypoxia response element (HRE) reporter gene by transfected EPAS1. Likewise, expression of a constitutively active MEK1 mimicked the effects of hypoxia on HRE reporter gene expression. However, pretreatment with PD98059 had no effect on EPAS1 phosphorylation during hypoxia, suggesting that MAPK targets other proteins that are critical for the trans-activation of EPAS1. We further show that hypoxia-induced trans-activation of EPAS1 is independent of Ras. Finally, pretreatment with calmodulin antagonists nearly completely blocked both the hypoxia-induced phosphorylation of MAPK and the EPAS1 trans-activation of HRE-Luc. These results demonstrate that the MAPK pathway is a critical mediator of EPAS1 activation and that activation of MAPK and EPAS1 occurs through a calmodulin- sensitive pathway and not through the GTPase, Ras. These results are the first to identify a specific signaling pathway involved in EPAS1 activation.",
author = "Conrad, {P. William} and Freeman, {Thomas L.} and Dana Beitner-Johnson and Millhorn, {David E.}",
year = "1999",
month = "11",
day = "19",
doi = "10.1074/jbc.274.47.33709",
language = "English (US)",
volume = "274",
pages = "33709--33713",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "47",

}

TY - JOUR

T1 - EPAS1 trans-activation during hypoxia requires p42/p44 MAPK

AU - Conrad, P. William

AU - Freeman, Thomas L.

AU - Beitner-Johnson, Dana

AU - Millhorn, David E.

PY - 1999/11/19

Y1 - 1999/11/19

N2 - Hypoxia is a common environmental stress that regulates gene expression and cell function. A number of hypoxia-regulated transcription factors have been identified and have been shown to play critical roles in mediating cellular responses to hypoxia. One of these is the endothelial PAS-domain protein 1 (EPAS1/HIF2-α/HLF/HRF). This protein is 48% homologous to hypoxia- inducible factor 1-α (HIF1-α). To date, virtually nothing is known about the signaling pathways that lead to either EPAS1 or HIF1-α activation. Here we show that EPAS1 is phosphorylated when PC12 cells are exposed to hypoxia and that p42/p44 MAPK is a critical mediator of EPAS1 activation. Pretreatment of PC12 cells with the MEK inhibitor, PD98059, completely blocked hypoxia-induced trans-activation of a hypoxia response element (HRE) reporter gene by transfected EPAS1. Likewise, expression of a constitutively active MEK1 mimicked the effects of hypoxia on HRE reporter gene expression. However, pretreatment with PD98059 had no effect on EPAS1 phosphorylation during hypoxia, suggesting that MAPK targets other proteins that are critical for the trans-activation of EPAS1. We further show that hypoxia-induced trans-activation of EPAS1 is independent of Ras. Finally, pretreatment with calmodulin antagonists nearly completely blocked both the hypoxia-induced phosphorylation of MAPK and the EPAS1 trans-activation of HRE-Luc. These results demonstrate that the MAPK pathway is a critical mediator of EPAS1 activation and that activation of MAPK and EPAS1 occurs through a calmodulin- sensitive pathway and not through the GTPase, Ras. These results are the first to identify a specific signaling pathway involved in EPAS1 activation.

AB - Hypoxia is a common environmental stress that regulates gene expression and cell function. A number of hypoxia-regulated transcription factors have been identified and have been shown to play critical roles in mediating cellular responses to hypoxia. One of these is the endothelial PAS-domain protein 1 (EPAS1/HIF2-α/HLF/HRF). This protein is 48% homologous to hypoxia- inducible factor 1-α (HIF1-α). To date, virtually nothing is known about the signaling pathways that lead to either EPAS1 or HIF1-α activation. Here we show that EPAS1 is phosphorylated when PC12 cells are exposed to hypoxia and that p42/p44 MAPK is a critical mediator of EPAS1 activation. Pretreatment of PC12 cells with the MEK inhibitor, PD98059, completely blocked hypoxia-induced trans-activation of a hypoxia response element (HRE) reporter gene by transfected EPAS1. Likewise, expression of a constitutively active MEK1 mimicked the effects of hypoxia on HRE reporter gene expression. However, pretreatment with PD98059 had no effect on EPAS1 phosphorylation during hypoxia, suggesting that MAPK targets other proteins that are critical for the trans-activation of EPAS1. We further show that hypoxia-induced trans-activation of EPAS1 is independent of Ras. Finally, pretreatment with calmodulin antagonists nearly completely blocked both the hypoxia-induced phosphorylation of MAPK and the EPAS1 trans-activation of HRE-Luc. These results demonstrate that the MAPK pathway is a critical mediator of EPAS1 activation and that activation of MAPK and EPAS1 occurs through a calmodulin- sensitive pathway and not through the GTPase, Ras. These results are the first to identify a specific signaling pathway involved in EPAS1 activation.

UR - http://www.scopus.com/inward/record.url?scp=0033584965&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033584965&partnerID=8YFLogxK

U2 - 10.1074/jbc.274.47.33709

DO - 10.1074/jbc.274.47.33709

M3 - Article

C2 - 10559262

AN - SCOPUS:0033584965

VL - 274

SP - 33709

EP - 33713

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 47

ER -