Cellular oxygen sensing: Importins and exportins are mediators of intracellular localisation of prolyl-4-hydroxylases PHD1 and PHD2

Steinhoff, Amrei; Pientka, Friederike Katharina; Möckel, Sylvia; Kettelhake, Antje; Hartmann, Enno; Köhler, M; Depping, Reinhard

doi:10.1016/j.bbrc.2009.07.090

Cited by 39 publications

(36 citation statements)

References 31 publications

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“…25 The P200 amino acid is located within the nuclear localization signal (NLS) implicated in the shuttling of PHD2 between the cytoplasm and the nucleus which plays a crucial role in HIF regulation. 26 We tested the shuttling of the P200Q mutation by immunofluorescence but we did not observe any impact of the mutation on the capacity of PHD2 to shuttle (data not shown).…”

Section: Resultsmentioning

confidence: 94%

Distinct deregulation of the hypoxia inducible factor by PHD2 mutants identified in germline DNA of patients with polycythemia

Ladroue

Hoogewijs²,

Gad³

et al. 2011

Haematologica

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BackgroundCongenital secondary erythrocytoses are due to deregulation of hypoxia inducible factor resulting in overproduction of erythropoietin. The most common germline mutation identified in the hypoxia signaling pathway is the Arginine 200-Tryptophan mutant of the von Hippel-Lindau tumor suppressor gene, resulting in Chuvash polycythemia. This mutant displays a weak deficiency in hypoxia inducible factor α regulation and does not promote tumorigenesis. Other von Hippel-Lindau mutants with more deleterious effects are responsible for von Hippel-Lindau disease, which is characterized by the development of multiple tumors. Recently, a few mutations in gene for the prolyl hydroxylase domain 2 protein (PHD2) have been reported in cases of congenital erythrocytosis not associated with tumor formation with the exception of one patient with a recurrent extra-adrenal paraganglioma. Design and MethodsFive PHD2 variants, four of which were novel, were identified in patients with erythrocytosis. These PHD2 variants were functionally analyzed and compared with the PHD2 mutant previously identified in a patient with polycythemia and paraganglioma. The capacity of PHD2 to regulate the activity, stability and hydroxylation of hypoxia inducible factor α was assessed using hypoxia-inducible reporter gene, one-hybrid and in vitro hydroxylation assays, respectively. ResultsThis functional comparative study showed that two categories of PHD2 mutants could be distinguished: one category with a weak deficiency in hypoxia inducible factor α regulation and a second one with a deleterious effect; the mutant implicated in tumor occurrence belongs to the second category. ConclusionsAs observed with germline von Hippel-Lindau mutations, there are functional differences between the PHD2 mutants with regards to hypoxia inducible factor regulation. PHD2 mutation carriers do, therefore, need careful medical follow-up, since some mutations must be considered as potential candidates for tumor predisposition.Key words: hypoxia inducible factor, hypoxia-inducible transcription factor, PHD2, erythrocytosis.Citation: Ladroue C, Hoogewijs D, Gad S, Carcenac R, Storti F, Barrois M, Gimenez-Roqueplo A-P, Leporrier M, Casadevall N, Hermine O, Kiladjian J-J, Baruchel A, Fakhoury F, Bressac-de Paillerets B, Feunteun J, Mazure N, Pouysségur J, Wenger RH, Richard S, and Gardie B. Distinct deregulation of the hypoxia inducible factor by PHD2 mutants identified in germline DNA of patients with polycythemia. Haematologica 2012;97(1):9-14.

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Section: Resultsmentioning

confidence: 94%

Distinct deregulation of the hypoxia inducible factor by PHD2 mutants identified in germline DNA of patients with polycythemia

Ladroue

Hoogewijs²,

Gad³

et al. 2011

Haematologica

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“…Investigations on intracellular localisation showed that PHD1 is present exclusively in the nucleus, PHD2 is mainly cytoplasmic and PHD3 is located both in the nucleus and the cytoplasm (Metzen et al, 2003;Steinhoff et al, 2009). In line with this data Wotzlaw and colleagues demonstrated that PHD1 and HIF-1a interact within the nuclear compartment (Wotzlaw et al, 2010).…”

Section: Introductionmentioning

confidence: 75%

“…The N-terminal part of the protein is essential for nuclear export as the deletion mutant PHD2D1-100 showed balanced nuclear and cytoplasmic localisation without inhibition of nuclear export (Steinhoff et al, 2009). Therefore, the EGFP-tagged N-terminal deletion mutant PHD2D6-20 was constructed and expressed in U2OS cells.…”

Section: Resultsmentioning

confidence: 99%

“…Moreover, we could demonstrate that nuclear import of PHD1 is importin-a/ b-dependent and relies on a nuclear localisation signal (NLS) (Steinhoff et al, 2009).…”

Section: Introductionmentioning

confidence: 92%

“…Despite similar in vitro enzymatic properties and the fact that overexpression of each PHD isoform suppresses HIF-dependent transcriptional activity differences among the three PHDs could be identified (Huang et al, 2002). Each PHD isoform is expressed in a tissue and cell type specific manner, has its distinct subcellular localisation pattern and shows differential activities towards HIF-1a and HIF-2a (Appelhoff et al, 2004;Metzen et al, 2003;Steinhoff et al, 2009). Inhibition of PHD2 by RNA interference was shown to be sufficient to upregulate HIF-1a in normoxia, suggesting PHD2 as the key regulator of HIF-1a in vivo (Appelhoff et al, 2004;Berra et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

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Oxygen sensing by Prolyl-4-Hydroxylase PHD2 within the nuclear compartment and the influence of compartimentalisation on HIF-1 signalling

Pientka

Schindler

et al. 2012

Journal of Cell Science

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SummaryHypoxia-inducible factors (HIFs) regulate more than 200 genes involved in cellular adaptation to reduced oxygen availability. HIFs are heterodimeric transcription factors that consist of one of three HIF-a subunits and a HIF-b subunit. Under normoxic conditions the HIFa subunit is hydroxylated by members of a family of prolyl-4-hydroxylase domain (PHD) proteins, PHD1, PHD2 and PHD3, resulting in recognition by von-Hippel-Lindau protein, ubiquitylation and proteasomal degradation. It has been suggested that PHD2 is the key regulator of HIF-1a stability in vivo. Previous studies on the intracellular distribution of PHD2 have provided evidence for a predominant cytoplasmic localisation but also nuclear activity of PHD2. Here, we investigated functional nuclear transport signals in PHD2 and identified amino acids 196-205 as having a crucial role in nuclear import, whereas amino acids 6-20 are important for nuclear export. Fluorescence resonance energy transfer (FRET) showed that an interaction between PHD2 and HIF-1a occurs in both the nuclear and cytoplasmic compartments. However, a PHD2 mutant that is restricted to the cytoplasm does not interact with HIF-1a and shows less prolyl hydroxylase activity for its target HIF-1a than wild-type PHD2 located in the nucleus. Here, we present a new model by which PHD2-mediated hydroxylation of HIF-1a predominantly occurs in the cell nucleus and is dependent on very dynamic subcellular trafficking of PHD2.

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Regulation of Oxygen Homeostasis by Prolyl Hydroxylase Domains

Takeda

Fong²

2010

Studies on Renal Disorders

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Cellular oxygen sensing: Importins and exportins are mediators of intracellular localisation of prolyl-4-hydroxylases PHD1 and PHD2

Cited by 39 publications

References 31 publications

Distinct deregulation of the hypoxia inducible factor by PHD2 mutants identified in germline DNA of patients with polycythemia

Distinct deregulation of the hypoxia inducible factor by PHD2 mutants identified in germline DNA of patients with polycythemia

Oxygen sensing by Prolyl-4-Hydroxylase PHD2 within the nuclear compartment and the influence of compartimentalisation on HIF-1 signalling

Regulation of Oxygen Homeostasis by Prolyl Hydroxylase Domains

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