Complex regulation of the transactivation function of hypoxia-inducible factor-1 alpha by direct interaction with two distinct domains of the CREB-binding protein/p300.
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 alpha. The C-terminal activation domain of HIF-1 alpha 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 alpha interacts with endogenous CBP and that this interaction facilitates its transactivation function. Our results show that interaction of HIF-1 alpha 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 alpha than of p53. Chromatin immunoprecipitation demonstrated that at 1% O(2) CBP is recruited to a HIF-1 alpha 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 alpha N-TAD and mediating the transactivation function of this domain. Thus, we suggest that in hypoxia HIF-1 alpha 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 alpha.
Ruas
Jorge L
JL
Department of Cell and Molecular Biology, Karolinska Institutet, von Eulers väg 3, S-171 77 Stockholm, Sweden.
Berchner-Pfannschmidt
Utta
U
Malik
Sohail
S
Gradin
Katarina
K
Fandrey
Joachim
J
Roeder
Robert G
RG
Pereira
Teresa
T
Poellinger
Lorenz
L
D015082:2,2'-Dipyridyl / Q000494:pharmacology; D002460:Cell Line; D002614:Chelating Agents / Q000494:pharmacology; D003545:Cysteine / Q000378:metabolism; D050881:E1A-Associated p300 Protein / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006639:Histidine / Q000378:metabolism; D006801:Humans; D000860:Hypoxia / Q000378:metabolism / Q000503:physiopathology*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007668:Kidney / Q000166:cytology; D016296:Mutagenesis; D054730:Protein Interaction Domains and Motifs / Q000187:drug effects / Q000502:physiology; D017434:Protein Structure, Tertiary; D015533:Transcriptional Activation / Q000502:physiology; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D050880:p300-CBP Transcription Factors / Q000737:chemistry / Q000378:metabolism*
D002614:Chelating Agents; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D016159:Tumor Suppressor Protein p53; D006639:Histidine; D015082:2,2'-Dipyridyl; D050881:E1A-Associated p300 Protein; D050880:p300-CBP Transcription Factors; D003545:Cysteine