Chelation of Cellular Calcium Modulates Hypoxia-inducible Gene Expression through Activation of Hypoxia-inducible Factor-1α

Berchner‐Pfannschmidt, Utta; Petrat, Frank; Doege, Kathrin; Trinidad, Buena; Freitag, Patrícia; Metzen, Eric; Groot, Herbert de; Fandrey, Joachim

doi:10.1074/jbc.m313995200

Cited by 58 publications

(44 citation statements)

References 51 publications

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“…It has been reported that chelation of intracellular Ca 2+ by bis-(O-aminophenoxy)-ethane-N,N,N′,N′-tetra-acetoxymethyl ester (BAPTA AM ) induces HIF-1α accumulation in the neuroblastoma cell line SH-SY5Y (Berchner-Pfannschmidt et al, 2004). In our work, an EGTA-induced reduction in the intracellular Ca 2+ levels and inhibition of TRPC6 suppressed the hypoxic accumulation of HIF-1α in human glioma cell lines.…”

Section: Discussionmentioning

confidence: 56%

Critical role of TRPC6 in maintaining the stability of HIF-1α in glioma cells under hypoxia

Wang

et al. 2015

Journal of Cell Science

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Hypoxia-inducible factor-1 (HIF-1) is a key transcription factor responsible for the expression of a broad range of genes that facilitate acclimatization to hypoxia. Its stability is predominantly controlled by rapid hydroxylation of two proline residues in its α-subunit. However, how the rapid hydroxylation of HIF-1α is regulated is not fully understood. Here, we report that transient receptor potential canonical (TRPC) 6 channels control hydroxylation and stability of HIF-1α in human glioma cells under hypoxia. TRPC6 was rapidly activated by IGF-1R-PLCγ-IP 3 R pathway upon hypoxia. Inhibition of TRPC6 enhanced the levels of α-ketoglutarate and promoted hydroxylation of HIF-1α to suppress HIF-1α accumulation without affecting its transcription or translation. Dimethyloxalylglycine N-(methoxyoxoacetyl)-glycine methyl ester (DMOG), an analog of α-ketoglutarate, reversed the inhibition of HIF-1α accumulation. Moreover, TRPC6 regulated GLUT1 (also known as SLC2A1) expression in a manner that was dependent on HIF-1α accumulation to affect glucose uptake during hypoxia. Our results suggest that TRPC6 regulates metabolism to affect HIF-1α stability and consequent glucose metabolism in human glioma cells under hypoxia.

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Section: Discussionmentioning

confidence: 56%

Critical role of TRPC6 in maintaining the stability of HIF-1α in glioma cells under hypoxia

Wang

et al. 2015

Journal of Cell Science

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“…Of course further experiments need to determine the temporally and spatially contribution of calpain-mediated destruction of HIF-1␣ relative to the proteasome system. Modulation of intracellular calcium is known to affect the expression of hypoxia-inducible genes and more recently, two reports provided evidence that lowering intracellular calcium by BAPTA-AM activates HIF-1 by attenuating hydroxylation of HIF-1␣ (Berchner-Pfannschmidt et al, 2004;Liu et al, 2004). Although the proposed mechanism will not be applicable to pVHL-deficient RCC4 cells, it supports the concept that calcium is involved in degradation of HIF-1␣.…”

Section: Discussionmentioning

confidence: 67%

Calpain Mediates a von Hippel-Lindau Protein–independent Destruction of Hypoxia-inducible Factor-1α

Zhou

Köhl

Herr

et al. 2006

MBoC

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Hypoxia-inducible factor 1 (HIF-1) is controlled through stability regulation of its alpha subunit, which is expressed under hypoxia but degraded under normoxia. Degradation of HIF-1␣ requires association of the von Hippel Lindau protein (pVHL) to provoke ubiquitination followed by proteasomal digestion. Besides hypoxia, nitric oxide (NO) stabilizes HIF-1␣ under normoxia but destabilizes the protein under hypoxia. To understand the role of NO under hypoxia we made use of pVHL-deficient renal carcinoma cells (RCC4) that show a high steady state HIF-1␣ expression under normoxia. Exposing RCC4 cells to hypoxia in combination with the NO donor DETA-NO (2,2-(hydroxynitrosohydrazono) bis-ethanimine), but not hypoxia or DETA-NO alone, decreased HIF-1␣ protein and attenuated HIF-1 transactivation. Mechanistically, we noticed a role of calpain because calpain inhibitors reversed HIF-1␣ degradation. Furthermore, chelating intracellular calcium attenuated HIF-1␣ destruction by hypoxia/DETA-NO, whereas a calcium increase was sufficient to lower the amount of HIF-1␣ even under normoxia. An active role of calpain in lowering HIF-1␣ amount was also evident in pVHL-containing human embryonic kidney cells when the calcium pump inhibitor thapsigargin reduced HIF-1␣ that was stabilized by the prolyl hydroxylase inhibitor dimethyloxalylglycine (DMOG). We conclude that calcium contributes to HIF-1␣ destruction involving the calpain system. INTRODUCTIONThe transcription factor hypoxia-inducible factor-1 (HIF-1) constitutes a central component in coordinating adaptive responses toward low oxygen availability, i.e., hypoxia. HIF-1 is a heterodimer composed of the 120-kDa HIF-1␣ subunit and the 91-94-kDa HIF-1␤ subunit (Semenza and Wang, 1992;Wang and Semenza, 1995). Although HIF-1␤ is constitutively expressed, oxygen facilitates continues destruction of HIF-1␣ via the 26S proteasome. This requires polyubiquitination by an E3-ubiquitin ligase complex that contains the von Hippel Lindau protein (pVHL; Kaelin, 2002;Bruick, 2003;Huang and Bunn, 2003;Pugh and Ratcliffe, 2003;Semenza, 2003). Ubiquitination and binding of pVHL to HIF-1␣ demands hydroxylation of Pro564 and/or Pro402 within the oxygen-dependent degradation domain (ODD) of HIF-1␣ (Ivan et al., 2001;Jaakkola et al., 2001;Masson et al., 2001). Hydroxylation is mediated by prolyl hydroxylases, also known as PH domain-containing enzymes (PHD, i.e., PHD1 to PHD4; Bruick and McKnight, 2001;Epstein et al., 2001;Oehme et al., 2002). In addition to regulating HIF-1␣ protein stability, oxygen affects the transcriptional activity of HIF-1, by regulating hydroxylation of a critical Asn803 residue within the C-terminal transactivation domain (CTAD) of HIF-1␣ (Lando et al., 2002). The asparagine hydroxylase known as FIH (factor inhibiting HIF; Mahon et al., 2001;Hewitson et al., 2002) renders CTAD unable to bind the coactivator p300/CBP. Thus, hypoxia attenuates Pro564/402 as well as Asn803 modifications, provoking protein stabilization and coactivator recruitment, which concomitantly results...

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“…FRET Experiments-HEK cells transfected with CFP-and YFP-fused expression plasmids were investigated in a chamber on the microscope stage (Luigs y Neumann) at normoxia at 37°C, as described previously (35)(36)(37)(38). 293 cells were grown on 35-mm dishes (WellcoDish) and transfected using FuGENE 6 according to the manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

“…Here we tested whether HIF-1␣ had the ability to interact with the CH3 region or other CBP domains. HIF-1␣ was 35 S-labeled by in vitro translation in rabbit reticulocyte lysate and incubated with different purified, bacterially expressed GST-fused CBP domains. As observed in Fig.…”

Section: Cbp Enhances the Transactivation Function Of A Hif-1␣ Mutantmentioning

confidence: 99%

Complex Regulation of the Transactivation Function of Hypoxia-inducible Factor-1α by Direct Interaction with Two Distinct Domains of the CREB-binding Protein/p300

Ruas

Berchner‐Pfannschmidt

Malik

et al. 2010

Journal of Biological Chemistry

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Activation of transcription in response to low oxygen tension is mediated by the hypoxia-inducible factor-1 (HIF-1). HIF-1 is a heterodimer of two proteins: aryl hydrocarbon receptor nuclear translocator and the oxygen-regulated HIF-1␣. The C-terminal activation domain of HIF-1␣ has been shown to interact with cysteine/ histidine-rich region 1 (CH1) of the coactivator CBP/p300 in a hypoxia-dependent manner. However, HIF forms lacking C-terminal activation domain (naturally occurring or genetically engineered) are still able to activate transcription of target genes in hypoxia. Here, we demonstrate that the N-terminal activation domain (N-TAD) of HIF-1␣ interacts with endogenous CBP and that this interaction facilitates its transactivation function. Our results show that interaction of HIF-1␣ N-TAD with CBP/p300 is mediated by the CH3 region of CBP known to interact with, among other factors, p53. Using fluorescence resonance energy transfer experiments, we demonstrate that N-TAD interacts with CH3 in vivo. Coimmunoprecipitation assays using endogenous proteins showed that immunoprecipitation of CBP in hypoxia results in the recovery of a larger fraction of HIF-1␣ than of p53. Chromatin immunoprecipitation demonstrated that at 1% O 2 CBP is recruited to a HIF-1␣ but not to a p53 target gene. Upon activation of both pathways, lower levels of chromatin-associated CBP were detected at either target gene promoter. These results identify CBP as the coactivator directly interacting with HIF-1␣ N-TAD and mediating the transactivation function of this domain. Thus, we suggest that in hypoxia HIF-1␣ is a major CBP-interacting transcription factor that may compete with other CBP-dependent factors, including p53, for limiting amounts of this coactivator, underscoring the complexity in the regulation of gene expression by HIF-1␣.

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Chelation of Cellular Calcium Modulates Hypoxia-inducible Gene Expression through Activation of Hypoxia-inducible Factor-1α

Cited by 58 publications

References 51 publications

Critical role of TRPC6 in maintaining the stability of HIF-1α in glioma cells under hypoxia

Critical role of TRPC6 in maintaining the stability of HIF-1α in glioma cells under hypoxia

Calpain Mediates a von Hippel-Lindau Protein–independent Destruction of Hypoxia-inducible Factor-1α

Complex Regulation of the Transactivation Function of Hypoxia-inducible Factor-1α by Direct Interaction with Two Distinct Domains of the CREB-binding Protein/p300

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