HIF-2α phosphorylation by CK1δ promotes erythropoietin secretion in liver cancer cells under hypoxia

Pangou, Evanthia; Befani, Christina; Mylonis, Ilias; Samiotaki, Martina; Panayotou, George; Simos, George; Liakos, Panagiotis

doi:10.1242/jcs.191395

Cited by 28 publications

(33 citation statements)

References 49 publications

(56 reference statements)

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“…Digested peptides were eluted from the filter twice with 100 µl water and dried in a vacuum centrifuge (Thermo Fisher Scientific, Waltham, MA). The samples were reconstituted with 20 μl solution of 2% (v/v) acetonitrile and 0.1% (v/v) formic acid, sonicated in a water bath for 3 min and analyzed with LC-MS/MS using an LTQ Orbitrap XL Mass spectrometer (Thermo Fisher Scientific) equipped with a nanospray source as previously described (Pangou et al, 2016). Protein identification was performed using Proteome Discoverer 1.4 software (Thermo Fisher Scientific) equipped with a SEQUEST HT search engine (Pangou et al, 2016).…”

Section: Tryptic Digestion Lc-ms/ms Analysis and Database Searchmentioning

confidence: 99%

“…The samples were reconstituted with 20 μl solution of 2% (v/v) acetonitrile and 0.1% (v/v) formic acid, sonicated in a water bath for 3 min and analyzed with LC-MS/MS using an LTQ Orbitrap XL Mass spectrometer (Thermo Fisher Scientific) equipped with a nanospray source as previously described (Pangou et al, 2016). Protein identification was performed using Proteome Discoverer 1.4 software (Thermo Fisher Scientific) equipped with a SEQUEST HT search engine (Pangou et al, 2016). Oxidation of methionine, deamidation of asparagine and glutamine, carbamidomethylation of cysteine and acetylation of the protein N-terminus were used as variable modifications.…”

Section: Tryptic Digestion Lc-ms/ms Analysis and Database Searchmentioning

confidence: 99%

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Mortalin-mediated and ERK-controlled targeting of HIF-1α to mitochondria confers resistance to apoptosis under hypoxia

Mylonis

Kourti

Samiotaki

et al. 2016

Journal of Cell Science

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Hypoxia inducible factor-1 (HIF-1) is the main transcriptional activator of the cellular response to hypoxia and an important target of anticancer therapy. Phosphorylation by ERK1 and/or ERK2 (MAPK3 and MAPK1, respectively; hereafter ERK) stimulates the transcriptional activity of HIF-1α by inhibiting its CRM1 (XPO1)-dependent nuclear export. Here, we demonstrate that phosphorylation by ERK also regulates the association of HIF-1α with a so-far-unknown interaction partner identified as mortalin (also known as GRP75 and HSPA9), which mediates non-genomic involvement of HIF-1α in apoptosis. Mortalin binds specifically to HIF-1α that lacks modification by ERK, and the HIF-1α-mortalin complex is localized outside the nucleus. Under hypoxia, mortalin mediates targeting of unmodified HIF-1α to the outer mitochondrial membrane, as well as association with VDAC1 and hexokinase II, which promotes production of a C-terminally truncated active form of VDAC1, denoted VDAC1-ΔC, and protection from apoptosis when ERK is inactivated. Under normoxia, transcriptionally inactive forms of unmodified HIF-1α or its C-terminal domain alone are also targeted to mitochondria, stimulate production of VDAC1-ΔC and increase resistance to etoposide-or doxorubicin-induced apoptosis. These findings reveal an ERK-controlled, unconventional and anti-apoptotic function of HIF-1α that might serve as an early protective mechanism upon oxygen limitation and promote cancer cell resistance to chemotherapy.

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Section: Tryptic Digestion Lc-ms/ms Analysis and Database Searchmentioning

confidence: 99%

Section: Tryptic Digestion Lc-ms/ms Analysis and Database Searchmentioning

confidence: 99%

Mortalin-mediated and ERK-controlled targeting of HIF-1α to mitochondria confers resistance to apoptosis under hypoxia

Mylonis

Kourti

Samiotaki

et al. 2016

Journal of Cell Science

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“…We have recently identified two residues of HIF-2α, S383 and T528 that are targeted by casein kinase 1δ (CK1δ, also known as CSNK1D). These modifications lead to efficient HIF-2α nuclear accumulation and full HIF-2 transcriptional activity (Pangou et al, 2016).…”

Section: Introductionmentioning

confidence: 99%

ERK1/2 phosphorylates HIF-2α and regulates its activity by controlling its CRM1-dependent nuclear shuttling

Gkotinakou

Befani

Simos

et al. 2019

Journal of Cell Science

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Hypoxia-inducible factor 2 (HIF-2) is a principal component of the cellular response to oxygen deprivation (hypoxia). Its inducible subunit, HIF-2α (also known as EPAS1), is controlled by oxygen-dependent as well as oxygen-independent mechanisms, such as phosphorylation. We show here that HIF-2α is phosphorylated under hypoxia (1% O 2) by extracellular signal-regulated protein kinases 1 and 2 (ERK1/2; also known as MAPK3 and MAPK1, respectively) at serine residue 672, as identified by in vitro phosphorylation assays. Mutation of this site to an alanine residue or inhibition of the ERK1/2 pathway decreases HIF-2 transcriptional activity and causes HIF-2α to mislocalize to the cytoplasm without changing its protein expression levels. Localization, reporter gene and immunoprecipitation experiments further show that HIF-2α associates with the exportin chromosomal maintenance 1 (CRM1, also known as XPO1) in a phosphorylation-sensitive manner and identify two critical leucine residues as part of an atypical CRM1-dependent nuclear export signal (NES) neighboring serine 672. Inhibition of CRM1 or mutation of these residues restores nuclear accumulation and activity of HIF-2α lacking the ERK1/2-mediated modification. In summary, we reveal a novel regulatory mechanism of HIF-2, involving ERK1/2-dependent phosphorylation of HIF-2α, which controls its nucleocytoplasmic shuttling and the HIF-2 transcriptional activity. This article has an associated First Person interview with the first author of the paper.

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“…pcDNA-flag-HIF1␣ and pcDNA-flag-HIF2␣ that were used for cotransfection experiments were described previously (23,24). pcDNA-His-SUMO-2 was previously described.…”

Section: Methodsmentioning

confidence: 99%

Hypoxia-induced Changes in SUMO Conjugation Affect Transcriptional Regulation Under Low Oxygen

Chachami

Stankovic‐Valentin

Karagiota

et al. 2019

Molecular & Cellular Proteomics

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HeLa cells grown under normoxia or hypoxia for 48h, were subjected to endogenous SUMO-immunoprecipitation in combination with quantitative mass spectrometry (SILAC) to gain insights into differences of the SUMO1 and SUMO2/3 proteome. Proteins whose SUMOylation changed without concomitant change in abundance were predominantly transcriptions factors. Particularly interesting was transcription factor TFAP2A (Activating enhancer binding Protein 2 alpha), whose sumoylation decreased on hypoxia. DeSUMOylation of TFAP2A enhanced transcriptional activity of HIF-1 under hypoxia contributing at the cellular response to low oxygen.

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HIF-2α phosphorylation by CK1δ promotes erythropoietin secretion in liver cancer cells under hypoxia

Cited by 28 publications

References 49 publications

Mortalin-mediated and ERK-controlled targeting of HIF-1α to mitochondria confers resistance to apoptosis under hypoxia

Mortalin-mediated and ERK-controlled targeting of HIF-1α to mitochondria confers resistance to apoptosis under hypoxia

ERK1/2 phosphorylates HIF-2α and regulates its activity by controlling its CRM1-dependent nuclear shuttling

Hypoxia-induced Changes in SUMO Conjugation Affect Transcriptional Regulation Under Low Oxygen

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