Hypoxia Induces c-fos Transcription via a Mitogen-activated Protein Kinase-dependent Pathway

Abstract

Hypoxia is a pathophysiological condition that occurs during injury, ischemia, and stroke. It is characterized by a decrease of reactive oxygen intermediates and a change of the intracellular redox level. In tumors hypoxia is regarded as a trigger for enhanced growth and metastasis. Here we report that in HeLa cells, hypoxic conditions induce the transcriptional activation of c-fos transcription via the serum response element. Mutations in the binding site for the ternary complex factor Elk-1 and the serum response factor abolished this induction, indicating that a ternary complex at the serum response element is necessary for the induction of the c-fos gene under hypoxia. The transcription factor Elk-1 was covalently modified by phosphorylation in response to hypoxia. Furthermore this hyperphosphorylation of Elk-1, the activation of mitogen-activated protein kinase (MAPK), and the induction of c-fos transcripts were blocked by PD98059, a specific inhibitor of mitogen-activated protein kinase kinase/extracellular signal-regulated protein kinase kinase 1. An in vitro kinase assay with Elk-1 as substrate showed that MAPK is activated under hypoxia. The activation of MAPK corresponds temporally with the phosphorylation and activation of Elk-1. Thus, a decrease of the intracellular reactive oxygen intermediate level by hypoxia induces c-fos via the MAPK pathway. These results suggest that the intracellular redox levels may be directly coupled to tumor growth, invasion, and metastasis via Elk-1-dependent induction of c-Fos controlled genes.The activation of c-fos by mitogens and changes of the intracellular redox level is mainly mediated by the serum response element (SRE).1 The SRE is a regulatory element found in many growth factor-regulated promotors that directs the rapid induction of gene expression (for review see Ref. 1). The best studied SRE is that of the c-fos gene. Two kinds of transcription factors are required for SRE activity: the ubiquitous transcription serum response factor (SRF) (2) and the ternary complex factors (TCFs), which form a ternary complex with the SRF. The human TCFs include Elk-1, SAP-1, and SAP-2 and constitute a subfamily within the Ets family of transcription factors (for review see Ref.3). These proteins need SRF to bind tightly to the SRE (4). The N-terminal domains of Elk-1 and SAP-1 mediate DNA contact and ternary complex formation. The transactivation domain at the C terminus contains several conserved MAPK phosphorylation sites. Growth factor-stimulated activation of the MAPK pathway results in phosphorylation of the Elk-1 C terminus (5), which then cooperates with the SRF C-terminal activation domain to activate transcription (6, 7).The transcription factor AP-1 is composed of members of the Fos family (c-Fos, Fos-B, Fra-1, and Fra-2) and the Jun family (c-Jun, JunB, and JunD) that form restricted homo-or heterodimers (for review see Ref. 8). With the exception of c-Jun homodimers, AP-1 is predominantly induced at a transcriptional level by novel synthesis of its subunits. Thi...

Hypoxia-associated Induction of Early Growth Response-1 Gene Expression

“…Hypoxia-mediated activation of transcription via Elk-1 in MPs, presumably in complex with serum response factor (43), is analogous to previous observations in HeLa cells concerning expression of c-fos (5,42). This pathway of hypoxia-associated induction of transcription at SREs may also explain, in part, SRE-dependent induction of the noninsulin-dependent glucose transporter (GLUT1) associated with oxygen deprivation and inhibitors of aerobic respiration (41).…”

“…Although HIF-1 is critical for restoring cellular homeostasis in hypoxia, biosynthetic events triggered by oxygen deprivation extend beyond HIF-1. For example, hypoxia leads to the induction of c-Fos and appearance of c-Jun/ c-Fos AP-1 heterodimers in cultured cells, potentially promoting expression of a range of genes, especially those involved in cell growth (5)(6)(7)(8). Oxygen deprivation also activates the transcription factor C/EBP␤ (9, 10), which, on binding to nuclear factor-interleukin 6 (NF-IL-6) motifs, results in expression of IL-6, a cytokine associated with acute inflammation.…”

Regulation of interleukin-8 expression by reduced oxygen pressure in human glioblastoma

“…The mitogen-activated protein kinase-dependent pathway is a candidate since it was recently shown to be involved in activating the c-fos gene in response to hypoxia in HeLa cells (Muller et al, 1997) Finally, to examine whether the in vitro di erences in mRNA inducing mechanisms between IL-8 and VEGF seen here had any correlates in vivo, we examined adjacent sections of human glioblastoma by in situ hybridization with probes for both mRNAs. Clear di erences were noted.…”

Novel chimeric gene promoters responsive to hypoxia and ionizing radiation

“…25 The same CArG sequence was shown to be responsible for HIF1-independent hypoxic activation of the c-fos gene, via a mitogen-activated protein kinase (MAPK) pathway involving the ternary complex factor Elk1. 26 However, when isolated from its natu-…”