Determining the hydroxylation status of NAA10-Trp38
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Q Exactive HF-X, Q Exactive HF
ORGANISM(S): Homo Sapiens (human)
SUBMITTER: 
Rasmus Ree
LAB HEAD: Thomas Arnesen
PROVIDER: PXD023655 | Pride | 2022-02-16
REPOSITORIES: Pride
Publications
Hydroxylation of the Acetyltransferase NAA10 Trp38 Is Not an Enzyme-Switch in Human Cells.
Ree Rasmus R Krogstad Karoline K McTiernan Nina N Jakobsson Magnus E ME Arnesen Thomas T
International journal of molecular sciences 20211030 21
NAA10 is a major <i>N</i>-terminal acetyltransferase (NAT) that catalyzes the cotranslational <i>N</i>-terminal (Nt-) acetylation of 40% of the human proteome. Several reports of lysine acetyltransferase (KAT) activity by NAA10 exist, but others have not been able to find any NAA10-derived KAT activity, the latter of which is supported by structural studies. The KAT activity of NAA10 towards hypoxia-inducible factor 1α (HIF-1α) was recently found to depend on the hydroxylation at Trp38 of NAA10 ...[more]
Similar Datasets
Project description:The hypoxia inducible factor (HIF) system orchestrates cellular responses to hypoxia in animals. HIF is an /-heterodimeric transcription factor that regulates the expression of hundreds of genes in a context dependent manner. A hypoxia-sensing component of the HIF system involves oxygen-dependent catalysis by the HIF hydroxylases; in humans there are three HIF prolyl hydroxylases (PHD1-3) and an asparaginyl hydroxylase (FIH). PHD catalysis regulates HIF levels and FIH catalysis regulates HIF activity. How differences in HIF hydroxylation status relate to variations in the induction of HIF target gene transcription is unknown. We report studies using small molecule inhibitors of the HIF hydroxylases to investigate the extent to which HIF target gene upregulation is induced by reduced PHD catalysis. The results reveal substantial differences in the role of prolyl- and asparaginyl-hydroxylation in regulating hypoxia responsive genes in cells. Selective PHD inhibitors with different structural scaffolds behave similarly. However, under the tested conditions, a broad-spectrum 2OG dioxygenase inhibitor is a better mimic of the transcriptional response to hypoxia than the selective PHD inhibitors, consistent with an important role for FIH in the hypoxic transcriptional response. Indeed, combined application of selective PHD and FIH inhibitors resulted in transcriptional induction of a subset of genes that were not fully responsive to PHD inhibition alone. Thus, for the therapeutic regulation of HIF target genes, it is important to consider both PHD and FIH activity, and in the case of some sets of target genes, simultaneous inhibition of the PHDs and FIH catalysis may be preferable.
2016-10-25 | E-MTAB-4264 | biostudies-arrayexpress
Project description:The hypoxia inducible factor (HIF) system orchestrates cellular responses to hypoxia in animals. HIF is an α/β-heterodimeric transcription factor that regulates the expression of hundreds of genes in a context dependent manner. A hypoxia-sensing component of the HIF system involves oxygen-dependent catalysis by the HIF hydroxylases; in humans there are three HIF prolyl hydroxylases (PHD1-3) and an asparaginyl hydroxylase (FIH). PHD catalysis regulates HIFα levels and FIH catalysis regulates HIF activity. How differences in HIFα hydroxylation status relate to variations in the induction of HIF target gene transcription is unknown. We report studies using small molecule inhibitors of the HIF hydroxylases to investigate the extent to which HIF target gene upregulation is induced by reduced PHD catalysis. The results reveal substantial differences in the role of prolyl- and asparaginyl-hydroxylation in regulating hypoxia responsive genes in cells. Selective PHD inhibitors with different structural scaffolds behave similarly. However, under the tested conditions, a broad-spectrum 2OG dioxygenase inhibitor is a better mimic of the transcriptional response to hypoxia than the selective PHD inhibitors, consistent with an important role for FIH in the hypoxic transcriptional response. Indeed, combined application of selective PHD and FIH inhibitors resulted in transcriptional induction of a subset of genes that were not fully responsive to PHD inhibition alone. Thus, for the therapeutic regulation of HIF target genes, it is important to consider both PHD and FIH activity, and in the case of some sets of target genes, simultaneous inhibition of the PHDs and FIH catalysis may be preferable.
2016-08-31 | E-MTAB-4300 | biostudies-arrayexpress
Project description:This a model from the article:
Hypoxia-dependent sequestration of an oxygen sensor by a widespread structural motif can shape the hypoxic response - a predictive kinetic model
Bernhard Schmierer, Béla Novák1 and Christopher J Schofield
BMC Systems Biology2010, 4:139
20955552,
Abstract:
Background
The activity of the heterodimeric transcription factor hypoxia inducible factor (HIF) is regulated by the post-translational, oxygen-dependent hydroxylation of its α-subunit by members of the prolyl hydroxylase domain (PHD or EGLN)-family and by factor inhibiting HIF (FIH). PHD-dependent hydroxylation targets HIFα for rapid proteasomal degradation; FIH-catalysed asparaginyl-hydroxylation of the C-terminal transactivation domain (CAD) of HIFα suppresses the CAD-dependent subset of the extensive transcriptional responses induced by HIF. FIH can also hydroxylate ankyrin-repeat domain (ARD) proteins, a large group of proteins which are functionally unrelated but share common structural features. Competition by ARD proteins for FIH is hypothesised to affect FIH activity towards HIFα; however the extent of this competition and its effect on the HIF-dependent hypoxic response are unknown.
Results
To analyse if and in which way the FIH/ARD protein interaction affects HIF-activity, we created a rate equation model. Our model predicts that an oxygen-regulated sequestration of FIH by ARD proteins significantly shapes the input/output characteristics of the HIF system. The FIH/ARD protein interaction is predicted to create an oxygen threshold for HIFα CAD-hydroxylation and to significantly sharpen the signal/response curves, which not only focuses HIFα CAD-hydroxylation into a defined range of oxygen tensions, but also makes the response ultrasensitive to varying oxygen tensions. Our model further suggests that the hydroxylation status of the ARD protein pool can encode the strength and the duration of a hypoxic episode, which may allow cells to memorise these features for a certain time period after reoxygenation.
Conclusions
The FIH/ARD protein interaction has the potential to contribute to oxygen-range finding, can sensitise the response to changes in oxygen levels, and can provide a memory of the strength and the duration of a hypoxic episode. These emergent properties are predicted to significantly shape the characteristics of HIF activity in animal cells. We argue that the FIH/ARD interaction should be taken into account in studies of the effect of pharmacological inhibition of the HIF-hydroxylases and propose that the interaction of a signalling sensor with a large group of proteins might be a general mechanism for the regulation of signalling pathways.
There are there models described in the paper. 1) Skeleton Model 1 (SKM1) - HIFα CAD-hydroxylation in the absence of the FIH/AR-interaction. 2) Skeleton Model 2 (SKM2) - FIG sequestration by ARD proteins and oxygen-dependent FIH-release. 3) Full Model (Fusion of SKM1 and SKM2) - the effects of the FIH/ARD proteins interaction on HIFα CAD-hydroxylation.
This model corresponds to the "Full Model" described in the paper. The model reproduces figure 5 of the publication.
This model originates from BioModels Database: A Database of Annotated Published Models (http://www.ebi.ac.uk/biomodels/). It is copyright (c) 2005-2011 The BioModels.net Team.
For more information see the terms of use.
To cite BioModels Database, please use: Li C, Donizelli M, Rodriguez N, Dharuri H, Endler L, Chelliah V, Li L, He E, Henry A, Stefan MI, Snoep JL, Hucka M, Le Novère N, Laibe C (2010) BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. BMC Syst Biol., 4:92.
2010-08-17 | BIOMD0000000300 | BioModels
Project description:The asparagine hydroxylase, factor inhibiting HIF (FIH) confers oxygen-dependence upon the hypoxia-inducible factor (HIF), a master regulator of the cellular adaptive response to hypoxia. Studies investigating whether asparagine hydroxylation is a general regulatory oxygen-dependent modification have identified multiple non-HIF targets for FIH. However the functional consequences of this outside of the HIF pathway remain unclear. Here, we demonstrate that the deubiquitinase ovarian tumor domain containing, ubiquitin aldehyde binding protein 1 (OTUB1) is a substrate for hydroxylation by endogenous FIH on N22. Mutation of N22 leads to a profound change in the interaction of OTUB1 with proteins important in cellular metabolism. Furthermore, mutant OTUB1 (lacking the hydroxylation site) impairs cellular metabolic processes when compared to wild type. Based on these data, we hypothesize that OTUB1 is a target for functional hydroxylation by FIH, and propose that this provides new insight into the regulation of cellular energy metabolism during hypoxia.
2016-01-11 | PXD002103 | Pride
Project description:The asparagine hydroxylase, factor inhibiting HIF (FIH) confers oxygen-dependence upon the hypoxia-inducible factor (HIF), a master regulator of the cellular adaptive response to hypoxia. Studies investigating whether asparagine hydroxylation is a general regulatory oxygen-dependent modification have identified multiple non-HIF targets for FIH. However the functional consequences of this outside of the HIF pathway remain unclear. Here, we demonstrate that the deubiquitinase ovarian tumor domain containing, ubiquitin aldehyde binding protein 1 (OTUB1) is a substrate for hydroxylation by endogenous FIH on N22. Mutation of N22 leads to a profound change in the interaction of OTUB1 with proteins important in cellular metabolism. Furthermore, mutant OTUB1 (lacking the hydroxylation site) impairs cellular metabolic processes when compared to wild type. Based on these data, we hypothesize that OTUB1 is a target for functional hydroxylation by FIH, and propose that this provides new insight into the regulation of cellular energy metabolism during hypoxia.
2017-03-28 | MSV000080695 | MassIVE
Project description:The aspariginyl-hydroxylase activity of Factor Inhibiting HIF (FIH) is a key regulator of the transcriptional activity of the hypoxia inducible factor. FIH also catalyses the hydroxylation of asparaginyl- and other-residues in ankyrin repeat domain (ARD) containing proteins, including Apoptosis stimulating of p53 (ASPP) protein family members. ASPP2 is reported to undergo a single FIH catalysed hydroxylation at Asn-986. We report biochemical and crystallographic evidence showing FIH can catalyse the unprecedented post-translational hydroxylation of both asparaginyl-residues in “VNVN” and related motifs of ankyrin repeat domains in apoptosis-stimulating of p53 (ASPP) proteins (i.e. ASPP1, ASPP2 and iASPP) and the related ASB11 and p18-INK4C proteins. The results extend the substrate scope of FIH catalysis and may have implications for its role in the hypoxic response and, ASPP protein function.
2022-08-11 | PXD017278 | Pride
Project description:Hypoxia occurs when tissue or cellular oxygen demand exceeds its supply and is a frequent condition in health and disease. Metazoans have developed a regulatory system that allows them to sense changes in oxygen levels in their microenvironment and adapt to them. The asparagine hydroxylase factor inhibiting HIF (FIH) regulates the transcriptional activity of the hypoxia-inducible factor (HIF), the master regulator of the cellular adaptive response to hypoxia [1, 2]. We recently identified the deubiquitinating enzyme ovarian tumour domain containing, ubiquitin aldehyde binding protein 1 (OTUB1) as novel bona fide FIH target protein with the hydroxylation occurring at the asparagine residue N22 [3, 4]. Surprisingly, in a follow-up investigation we observed a SDS-PAGE resistant interaction between FIH and OTUB1, resulting in a FIH-OTUB1 heterodimer (HD). Further analysis of the FIH-OTUB1 HD demonstrated that it is not linked by a disulfide bond, oxyester or thioester but likely through an amide bond. Interestingly, mutation of the hydroxylation acceptor site N22 in OTUB1 and genetic and pharmacologic inhibition of FIH prevented the formation of the heterodimer, demonstrating that HD formation is a consequence of FIH activity. To investigate if FIH-dependent stable protein complex formation was exclusive for OTUB1, we carried out a mass spectrometry (MS)-based FIH interactome analyses with denatured and non-denatured cell lysates (with or without SDS treatment and boiling). The results indicate the existence of further novel denaturing condition resistant protein complexes. 1. Mahon, P.C., K. Hirota, and G.L. Semenza, FIH-1: a novel protein that interacts with HIF-1alpha and VHL to mediate repression of HIF-1 transcriptional activity. Genes & development, 2001. 15(20): p. 2675-86. 2. Lando, D., et al., FIH-1 is an asparaginyl hydroxylase enzyme that regulates the transcriptional activity of hypoxia-inducible factor. Genes & development, 2002. 16(12): p. 1466-71. 3. Scholz, C.C., et al., Regulation of IL-1beta-induced NF-kappaB by hydroxylases links key hypoxic and inflammatory signaling pathways. Proc Natl Acad Sci U S A, 2013. 110(46): p. 18490-5. 4. Scholz, C.C., et al., FIH Regulates Cellular Metabolism through Hydroxylation of the Deubiquitinase OTUB1. PLoS biology, 2016. 14(1): p. e1002347.
2019-07-09 | PXD011252 | Pride
Project description:Hypoxia Inducible Factor (HIF) prolyl hydroxylase domain (PHD) enzymes catalyse the posttranslational hydroxylation of conserved prolyl residues in the alpha-subunit of the HIF transcription factor. These modifications, which promote the degradation of HIF-alpha subunits by the pVHL E3 ligase complex and impart oxygen-dependent regulation of the HIF transcriptional response, have been extensively characterised at the molecular, cellular and organismal level. Since the discovery of the PHDs, a range of less well-characterised non-HIF substrates have been reported with the potential to confer oxygen-sensitivity to a diverse range of cellular processes. We sought to systematically compare all of the reported non-HIF substrates for their ability to support PHD-catalysed hydroxylation. We performed a comprehensive series of in vitro hydroxylation assays reacting synthetic peptides and full-length protein substrates generated by in vitro transcription and translation with purified recombinant enzyme preparations. Prolyl hydroxylation was assayed directly by mass spectrometry and radiochemical assay for hydroxyproline. Both methods enabled quantitative appraisal of enzyme-catalysed hydroxylation, with liquid chromatography mass spectrometry methods employing NMR-quantified peptide standards to calibrate retention time signatures and determine ionisation efficiencies for each target peptide. Using these approaches we assayed hydroxylation on 23 different proteins. Surprisingly, we did not detect measurable hydroxylation on any of the reported non-HIF substrates using either method. In contrast, control assays with HIF1-alpha substrate supported high stoichiometry (typically >90%) hydroxylation. Our findings suggest that PHD substrates are more restricted than has been reported.
2019-08-30 | PXD013112 | Pride
Project description:Identification of IRF3 and HIF-1α hydroxylation site(s) by mass spectrometry
2024-01-18 | PXD048667 | iProX
Project description:Clioquinol (CQ) has been identified as a hypoxic mimicker to activate hypoxia-inducible factor-1α (HIF-1α) by inhibiting HIF-1a specific asparaginyl hypoxylase (Factor inhibiting HIF-1, FIH-1). We previously showed that hypoxia coordinately regulate induction or repression of hypoxia-responsive genes by altering different types of methylations (H3K4me3, H3K9me3, and H3K27me3) at gene level. Here, we investigated CQ-mediated regulation of the expression of hypoxia-responsive genes and whether CQ had a similar effect on histone methylation and transcription of target genes under hypoxia.
2022-05-25 | GSE176557 | GEO