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HIF Prolyl-4-Hydroxylase PHD2 protein abundance depends on integral membrane-anchoring of FKBP38


Barth, S; Edlich, F; Berchner-Pfannschmidt, U; Gneuss, S; Jahreis, G; Hasgall, P A; Fandrey, J; Wenger, R H; Camenisch, G (2009). HIF Prolyl-4-Hydroxylase PHD2 protein abundance depends on integral membrane-anchoring of FKBP38. Journal of Biological Chemistry, 284(34):23046-23058.

Abstract

Prolyl-4-hydroxylase domain proteins (PHDs)
are 2-oxoglutarate and dioxygen-dependent
enzymes that mediate the rapid destruction of
hypoxia-inducible factor (HIF) α 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 HIFα
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 peptidyl prolyl cis/trans isomerase (PPIase)-
independent manner. Using peptide array
binding assays, fluorescence spectroscopy and
fluorescence resonance energy transfer (FRET)
analysis, we define a minimal linear glutamaterich
PHD2 binding domain in the N-terminal
part of FKBP38 and show that this domain
forms a high affinity complex with PHD2. Vice
versa, PHD2 interacted with a non-linear Nterminal
motif containing the MYND-type Zn2+
finger domain with FKBP38. Biochemical
fractionation and immunofluorescence analysis
demonstrated that PHD2 subcellular
localization overlapped with FKBP38 in the
endoplasmic reticulum (ER) 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
membrane-anchored FKBP38 localization
mediated by the C-terminal transmembrane
domain. Mechanistically, our data indicate that
PHD2 protein stability is regulated by an
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 active prolyl-4-
hydroxylase.Prolyl-4-hydroxylase domain proteins (PHDs)
are 2-oxoglutarate and dioxygen-dependent
enzymes that mediate the rapid destruction of
hypoxia-inducible factor (HIF) α 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 HIFα
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 peptidyl prolyl cis/trans isomerase (PPIase)-
independent manner. Using peptide array
binding assays, fluorescence spectroscopy and
fluorescence resonance energy transfer (FRET)
analysis, we define a minimal linear glutamaterich
PHD2 binding domain in the N-terminal
part of FKBP38 and show that this domain
forms a high affinity complex with PHD2. Vice
versa, PHD2 interacted with a non-linear Nterminal
motif containing the MYND-type Zn2+
finger domain with FKBP38. Biochemical
fractionation and immunofluorescence analysis
demonstrated that PHD2 subcellular
localization overlapped with FKBP38 in the
endoplasmic reticulum (ER) 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
membrane-anchored FKBP38 localization
mediated by the C-terminal transmembrane
domain. Mechanistically, our data indicate that
PHD2 protein stability is regulated by an
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 active prolyl-4-
hydroxylase.

Prolyl-4-hydroxylase domain proteins (PHDs)
are 2-oxoglutarate and dioxygen-dependent
enzymes that mediate the rapid destruction of
hypoxia-inducible factor (HIF) α 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 HIFα
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 peptidyl prolyl cis/trans isomerase (PPIase)-
independent manner. Using peptide array
binding assays, fluorescence spectroscopy and
fluorescence resonance energy transfer (FRET)
analysis, we define a minimal linear glutamaterich
PHD2 binding domain in the N-terminal
part of FKBP38 and show that this domain
forms a high affinity complex with PHD2. Vice
versa, PHD2 interacted with a non-linear Nterminal
motif containing the MYND-type Zn2+
finger domain with FKBP38. Biochemical
fractionation and immunofluorescence analysis
demonstrated that PHD2 subcellular
localization overlapped with FKBP38 in the
endoplasmic reticulum (ER) 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
membrane-anchored FKBP38 localization
mediated by the C-terminal transmembrane
domain. Mechanistically, our data indicate that
PHD2 protein stability is regulated by an
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 active prolyl-4-
hydroxylase.Prolyl-4-hydroxylase domain proteins (PHDs)
are 2-oxoglutarate and dioxygen-dependent
enzymes that mediate the rapid destruction of
hypoxia-inducible factor (HIF) α 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 HIFα
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 peptidyl prolyl cis/trans isomerase (PPIase)-
independent manner. Using peptide array
binding assays, fluorescence spectroscopy and
fluorescence resonance energy transfer (FRET)
analysis, we define a minimal linear glutamaterich
PHD2 binding domain in the N-terminal
part of FKBP38 and show that this domain
forms a high affinity complex with PHD2. Vice
versa, PHD2 interacted with a non-linear Nterminal
motif containing the MYND-type Zn2+
finger domain with FKBP38. Biochemical
fractionation and immunofluorescence analysis
demonstrated that PHD2 subcellular
localization overlapped with FKBP38 in the
endoplasmic reticulum (ER) 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
membrane-anchored FKBP38 localization
mediated by the C-terminal transmembrane
domain. Mechanistically, our data indicate that
PHD2 protein stability is regulated by an
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 active prolyl-4-
hydroxylase.

Citations

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Center for Integrative Human Physiology
04 Faculty of Medicine > Institute of Physiology
07 Faculty of Science > Institute of Physiology
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:22 August 2009
Deposited On:13 Jul 2009 05:10
Last Modified:26 Aug 2016 07:32
Publisher:American Society for Biochemistry and Molecular Biology
ISSN:0021-9258
Funders:Forschungskredit der Universität Zürich, Edoardo R., Giovanni, Giuseppe und Chiarina Sassella-Stiftung, Zürich, Hartmann Müller-Stiftung für Medizinische Forschung, Zürich, Krebsliga des Kantons Zürich, Olga Mayenfisch Stiftung, Zürich, 6th Framework Programme of the European Commission/SBF, Swiss National Foundation, Deutsche Forschungsgemeinschaft
Additional Information:This research was originally published in Barth, S; Edlich, F; Berchner-Pfannschmidt, U; Gneuss, S; Jahreis, G; Hasgall, P A; Fandrey, J; Wenger, R H; Camenisch, G (2009). HIF Prolyl-4-Hydroxylase PHD2 protein abundance depends on integral membrane-anchoring of FKBP38. Journal of Biological Chemistry, 284(34):23046-23058. © the American Society for Biochemistry and Molecular Biology.
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1074/jbc.M109.032631
PubMed ID:19546213
Other Identification Number:M109.032631
Permanent URL: https://doi.org/10.5167/uzh-19586

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