Imaging of the hypoxia-inducible factor pathway: insights into oxygen sensing

Berchner‐Pfannschmidt, Utta; Frede, Stilla; Wotzlaw, Christoph; Fandrey, Joachim

doi:10.1183/09031936.00013408

Cited by 43 publications

(39 citation statements)

References 54 publications

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“…The measurements resulted in a K D value of 1.48 Ϯ 0.15 M. Isothermal titration calorimetry measurements characterized the interaction between PHD2 and the N-terminal extension of FKBP38 as a 1:1 complex with a K D of 895 Ϯ 148 nM (data not shown). The interaction between PHD2 and FKBP38 [37][38][39][40][41][42][43][44][45][46][47][48][49][50][51][52][53][54][55][56] was measured in parallel by isothermal titration calorimetry resulting in a K D of 1.28 Ϯ 0.19 M, which is similar to the result of the fluorescence titration curve.…”

Section: Biochemical Characterization Of the Phd2/fkbp38supporting

confidence: 66%

Hypoxia-inducible Factor Prolyl-4-hydroxylase PHD2 Protein Abundance Depends on Integral Membrane Anchoring of FKBP38

Barth

Edlich

Berchner‐Pfannschmidt

et al. 2009

Journal of Biological Chemistry

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Prolyl-4-hydroxylase domain (PHD) proteins are 2-oxoglutarate and dioxygen-dependent enzymes that mediate the rapid destruction of hypoxia-inducible factor ␣ subunits. Whereas PHD1 and PHD3 proteolysis has been shown to be regulated by Siah2 ubiquitin E3 ligase-mediated polyubiquitylation and proteasomal destruction, protein regulation of the main oxygen sensor responsible for hypoxia-inducible factor ␣ regulation, PHD2, remained unknown. We recently reported that the FK506-binding protein (FKBP) 38 specifically interacts with PHD2 and determines PHD2 protein stability in a peptidylprolyl cis-trans isomerase-independent manner. Using peptide array binding assays, fluorescence spectroscopy, and fluorescence resonance energy transfer analysis, we defined a minimal linear glutamate-rich PHD2 binding domain in the N-terminal part of FKBP38 and showed that this domain forms a high affinity complex with PHD2. Vice versa, PHD2 interacted with a nonlinear N-terminal motif containing the MYND (myeloid, Nervy, and DEAF-1)-type Zn 2؉ finger domain with FKBP38. Biochemical fractionation and immunofluorescence analysis demonstrated that PHD2 subcellular localization overlapped with FKBP38 in the endoplasmic reticulum and mitochondria. An additional fraction of PHD2 was found in the cytoplasm. In cellulo PHD2/FKBP38 association, as well as regulation of PHD2 protein abundance by FKBP38, is dependent on membraneanchored FKBP38 localization mediated by the C-terminal transmembrane domain. Mechanistically our data indicate that PHD2 protein stability is regulated by a ubiquitin-independent proteasomal pathway involving FKBP38 as adaptor protein that mediates proteasomal interaction. We hypothesize that FKBP38-bound PHD2 is constantly degraded whereas cytosolic PHD2 is stable and able to function as an active prolyl-4-hydroxylase.The heterodimeric ␣/␤ transcription factor complexes of hypoxia-inducible factors (HIFs) 4 are central regulators of the cellular, local, and systemic response to reduced oxygen partial pressure (pO 2 ) (1, 2). Under normoxic conditions, two highly conserved prolyl residues within the oxygen-dependent degradation domain of HIF␣ subunits are hydroxylated by members of the prolyl-4-hydroxylase domain (PHD) family (also called egg laying-defective nine homolog (EGLN) or HIF prolyl hydroxylase) (3-5). Hydroxylated prolines are then bound by an E3 ubiquitin ligase complex containing the von Hippel-Lindau tumor suppressor protein (pVHL) as recognition subunit, mediating polyubiquitylation and proteasomal degradation of HIF␣ subunits (6 -8). In addition, factor inhibiting HIF hydroxylates under normoxic conditions an asparaginyl residue in the C-terminal transcriptional transactivation domain of HIF␣ subunits, preventing the association with the CH1 domain of the p300 and cAMP-responsive element-binding protein-binding protein (CBP) co-activators (9, 10). PHDs and factor inhibiting HIF belong to the 2-oxoglutarate-and irondependent dioxygenase superfamily and act as cellular oxygen sensors by correlating the ...

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Section: Biochemical Characterization Of the Phd2/fkbp38supporting

confidence: 66%

Hypoxia-inducible Factor Prolyl-4-hydroxylase PHD2 Protein Abundance Depends on Integral Membrane Anchoring of FKBP38

Barth

Edlich

Berchner‐Pfannschmidt

et al. 2009

Journal of Biological Chemistry

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“…It is the current opinion that hypoxia has no effect on the levels of HIF-1β (40,41). So we and most other researchers only focus our study on the oxygen-regulated HIF-1α subunit.…”

Section: Discussionmentioning

confidence: 97%

Oligomer procyanidins (F2) isolated from grape seeds inhibits tumor angiogenesis and cell invasion by targeting HIF-1α in vitro

Zheng

Yang

Hou

et al. 2014

International Journal of Oncology

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Abstract. Overexpression of hypoxia-inducible factor-1 (HIF-1) α, a transcription factor which immortalizes tumors by inducing expression of the genes involved in cell survival, migration and angiogenesis, is closely associated with poor prognosis, increased risk of metastasis and increased mortality. Oligomer procyanidins (F2), a natural fraction from grape seeds, has been demonstrated to have antioxidant and antitumor activities, however the antitumor effect of F2 targeting HIF-1α remains unknown. The present study showed that F2 markedly decreased HIF-1α and the expression of its target genes in cancer cells through inactivating the EGFR-PI3K-AKT-mTOR and MAPK-ERK1/2 pathways. Moreover, F2 suppressed vascular endothelial growth factor (VEGF) and matrix metalloproteinases (MMP)-2 expressions, followed by the inhibition of tumor angiogenesis and cell invasion in a HIF-1α-dependent manner. Collectively, these findings indicate that the antitumor effect of F2 is, at least in part, mediated by suppressing HIF-1α-dependent pathway, and suggest that F2 may be a potentially useful agent for treatment of human cancer.

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“…The α-subunit is hydroxylated at conserved prolyl and asparaginyl residues and targeted for degradation by the von Hippel-Lindau (VHL) ubiquitin E3 ligase complex under normal oxygen conditions. In response to low levels of oxygen, HIF-1α becomes activated and dimerizes with a constitutively expressed HIF-1β, and the complex binds to cis-acting hypoxia response elements in the promoter of target genes, which eventually regulates many cellular activities involved in oxygen homeostasis [3,4,11,[29][30][31]. Consistent with this, Harvey [7,32] suggested that HIFs may be involved in molecular mechanisms which respond to changes in oxygen status during embryo development by expressing genes of HIF-1α, -2α, and -1β subunits in mouse and bovine blastocysts.…”

Section: Discussionmentioning

confidence: 99%

“…In order for HIF-1α to act as a transcription factor, it should be observed in the nucleus. HIF-1α is translocated into the nucleus and dimerized with HIF-1β, and then the HIF-1 complexes bind to DNA regulatory sequences of HIF-1 target genes [29]. However, in this study HIF-1α protein was determined to be localized only in the cytoplasm, which is consistent with the previous study conducted by Thompson et al [33] that HIF-1α was detected in the cytoplasm of mouse blastocysts when cultured in low (2 %) oxygen concentration during compaction and blastulation.…”

Section: Discussionmentioning

confidence: 99%

Effects of oxygen tension and IGF-I on HIF-1α protein expression in mouse blastocysts

Yoon

Juhn

et al. 2012

J Assist Reprod Genet

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Purpose Hypoxia inducible factors (HIFs) are key regulators of oxygen homeostasis in response to reduced oxygenation in somatic cells. In addition, HIF-1α protein can be also induced by insulin-like growth factor I (IGF-I) treatment in various cell lines under normoxic condition. However, the expression and function of HIF-1α in embryogenesis are still unclear. Therefore, the objectives of this study were to examine the expression of HIF-1α in mouse blastocysts cultured under hypoxic and normoxic conditions, and to determine whether oxygen tension and IGF-I influence embryonic development through stimulation of HIF-1α expression. Methods Mouse embryos were cultured from the 1-cell to blastocyst stage under 5 % or 20 % O 2 in both the absence and presence of IGF-I. Results The embryonic development rates to the blastocyst stage were not affected by oxygen tension or IGF-I treatment. HIF-1α protein was localized to the cytoplasm of blastocysts, and its levels were independent of oxygen concentration or IGF-I treatment. Blastocysts cultured under 5 % O 2 exhibited significantly higher total cell numbers (83.4±18.1) and lower apoptotic index (3.7±1.5) than those cultured under 20 % O 2 (67.4±15.6) (6.9±3.5) (P<0.05). IGF-I reduced the apoptotic index in both oxygen conditions, but a significant decrease was detected in the 20 % O 2 group. Conclusions HIF-1α may not be a major mediator that responds to change in oxygen tension within blastocysts, inconsistent with that of somatic cells. Supplementation of culture media with IGF-I has been shown to promote embryo development by an anti-apoptotic effect, instead of increasing HIF-1α protein expression.

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Imaging of the hypoxia-inducible factor pathway: insights into oxygen sensing

Cited by 43 publications

References 54 publications

Hypoxia-inducible Factor Prolyl-4-hydroxylase PHD2 Protein Abundance Depends on Integral Membrane Anchoring of FKBP38

Hypoxia-inducible Factor Prolyl-4-hydroxylase PHD2 Protein Abundance Depends on Integral Membrane Anchoring of FKBP38

Oligomer procyanidins (F2) isolated from grape seeds inhibits tumor angiogenesis and cell invasion by targeting HIF-1α in vitro

Effects of oxygen tension and IGF-I on HIF-1α protein expression in mouse blastocysts

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