The LIMD1 protein bridges an association between the prolyl hydroxylases and VHL to repress HIF-1 activity

Katherine S Bridge; Daniel E Foxler; Victoria James; Thomas M Webb; Maureen Mee; Sybil C K Wong; Yunfeng Feng; Dumitru Constantin-Teodosiu; Thorgunnur Eyfjord Petursdottir; Johannes Bjornsson; Sigurdur Ingvarsson; Peter J Ratcliffe; Gregory D Longmore; Tyson V Sharp

doi:10.1038/ncb2424

The LIMD1 protein bridges an association between the prolyl hydroxylases and VHL to repress HIF-1 activity

Katherine S Bridge, Daniel E Foxler, Victoria James, Thomas M Webb, Maureen Mee, Sybil C K Wong, Yunfeng Feng, Dumitru Constantin-Teodosiu, Thorgunnur Eyfjord Petursdottir, Johannes Bjornsson, Sigurdur Ingvarsson, Peter J Ratcliffe, Gregory D Longmore, Tyson V Sharp

Biology

Research output: Contribution to journal › Article › peer-review

Abstract

There are three prolyl hydroxylases (PHD1, 2 and 3) that regulate the hypoxia-inducible factors (HIFs), the master transcriptional regulators that respond to changes in intracellular O(2) tension. In high O(2) tension (normoxia) the PHDs hydroxylate two conserved proline residues on HIF-1α, which leads to binding of the von Hippel-Lindau (VHL) tumour suppressor, the recognition component of a ubiquitin-ligase complex, initiating HIF-1α ubiquitylation and degradation. However, it is not known whether PHDs and VHL act separately to exert their enzymatic activities on HIF-1α or as a multiprotein complex. Here we show that the tumour suppressor protein LIMD1 (LIM domain-containing protein) acts as a molecular scaffold, simultaneously binding the PHDs and VHL, thereby assembling a PHD-LIMD1-VHL protein complex and creating an enzymatic niche that enables efficient degradation of HIF-1α. Depletion of endogenous LIMD1 increases HIF-1α levels and transcriptional activity in both normoxia and hypoxia. Conversely, LIMD1 expression downregulates HIF-1 transcriptional activity in a manner depending on PHD and 26S proteasome activities. LIMD1 family member proteins Ajuba and WTIP also bind to VHL and PHDs 1 and 3, indicating that these LIM domain-containing proteins represent a previously unrecognized group of hypoxic regulators.

Original language	English
Pages (from-to)	201-8
Number of pages	8
Journal	Nature Cell Biology
Volume	14
Issue number	2
DOIs	https://doi.org/10.1038/ncb2424
Publication status	Published - 29 Jan 2012

Keywords

Cell Hypoxia
Cell Line, Tumor
HEK293 Cells
HeLa Cells
Humans
Hydroxylation
Hypoxia-Inducible Factor 1, alpha Subunit/genetics
Hypoxia-Inducible Factor-Proline Dioxygenases
Immunoblotting
Immunoprecipitation
Intracellular Signaling Peptides and Proteins/genetics
LIM Domain Proteins/genetics
Models, Biological
Polyubiquitin/metabolism
Procollagen-Proline Dioxygenase/genetics
Proteasome Endopeptidase Complex/metabolism
Protein Binding
RNA Interference
Transfection
Two-Hybrid System Techniques
Ubiquitination
Von Hippel-Lindau Tumor Suppressor Protein/genetics

Access to Document

10.1038/ncb2424

Cite this

Bridge, K. S., Foxler, D. E., James, V., Webb, T. M., Mee, M., Wong, S. C. K., Feng, Y., Constantin-Teodosiu, D., Petursdottir, T. E., Bjornsson, J., Ingvarsson, S., Ratcliffe, P. J., Longmore, G. D., & Sharp, T. V. (2012). The LIMD1 protein bridges an association between the prolyl hydroxylases and VHL to repress HIF-1 activity. Nature Cell Biology, 14(2), 201-8. https://doi.org/10.1038/ncb2424

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abstract = "There are three prolyl hydroxylases (PHD1, 2 and 3) that regulate the hypoxia-inducible factors (HIFs), the master transcriptional regulators that respond to changes in intracellular O(2) tension. In high O(2) tension (normoxia) the PHDs hydroxylate two conserved proline residues on HIF-1α, which leads to binding of the von Hippel-Lindau (VHL) tumour suppressor, the recognition component of a ubiquitin-ligase complex, initiating HIF-1α ubiquitylation and degradation. However, it is not known whether PHDs and VHL act separately to exert their enzymatic activities on HIF-1α or as a multiprotein complex. Here we show that the tumour suppressor protein LIMD1 (LIM domain-containing protein) acts as a molecular scaffold, simultaneously binding the PHDs and VHL, thereby assembling a PHD-LIMD1-VHL protein complex and creating an enzymatic niche that enables efficient degradation of HIF-1α. Depletion of endogenous LIMD1 increases HIF-1α levels and transcriptional activity in both normoxia and hypoxia. Conversely, LIMD1 expression downregulates HIF-1 transcriptional activity in a manner depending on PHD and 26S proteasome activities. LIMD1 family member proteins Ajuba and WTIP also bind to VHL and PHDs 1 and 3, indicating that these LIM domain-containing proteins represent a previously unrecognized group of hypoxic regulators.",

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author = "Bridge, {Katherine S} and Foxler, {Daniel E} and Victoria James and Webb, {Thomas M} and Maureen Mee and Wong, {Sybil C K} and Yunfeng Feng and Dumitru Constantin-Teodosiu and Petursdottir, {Thorgunnur Eyfjord} and Johannes Bjornsson and Sigurdur Ingvarsson and Ratcliffe, {Peter J} and Longmore, {Gregory D} and Sharp, {Tyson V}",

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Bridge, KS, Foxler, DE, James, V, Webb, TM, Mee, M, Wong, SCK, Feng, Y, Constantin-Teodosiu, D, Petursdottir, TE, Bjornsson, J, Ingvarsson, S, Ratcliffe, PJ, Longmore, GD & Sharp, TV 2012, 'The LIMD1 protein bridges an association between the prolyl hydroxylases and VHL to repress HIF-1 activity', Nature Cell Biology, vol. 14, no. 2, pp. 201-8. https://doi.org/10.1038/ncb2424

TY - JOUR

T1 - The LIMD1 protein bridges an association between the prolyl hydroxylases and VHL to repress HIF-1 activity

AU - Bridge, Katherine S

AU - Foxler, Daniel E

AU - James, Victoria

AU - Webb, Thomas M

AU - Mee, Maureen

AU - Wong, Sybil C K

AU - Feng, Yunfeng

AU - Constantin-Teodosiu, Dumitru

AU - Petursdottir, Thorgunnur Eyfjord

AU - Bjornsson, Johannes

AU - Ingvarsson, Sigurdur

AU - Ratcliffe, Peter J

AU - Longmore, Gregory D

AU - Sharp, Tyson V

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N2 - There are three prolyl hydroxylases (PHD1, 2 and 3) that regulate the hypoxia-inducible factors (HIFs), the master transcriptional regulators that respond to changes in intracellular O(2) tension. In high O(2) tension (normoxia) the PHDs hydroxylate two conserved proline residues on HIF-1α, which leads to binding of the von Hippel-Lindau (VHL) tumour suppressor, the recognition component of a ubiquitin-ligase complex, initiating HIF-1α ubiquitylation and degradation. However, it is not known whether PHDs and VHL act separately to exert their enzymatic activities on HIF-1α or as a multiprotein complex. Here we show that the tumour suppressor protein LIMD1 (LIM domain-containing protein) acts as a molecular scaffold, simultaneously binding the PHDs and VHL, thereby assembling a PHD-LIMD1-VHL protein complex and creating an enzymatic niche that enables efficient degradation of HIF-1α. Depletion of endogenous LIMD1 increases HIF-1α levels and transcriptional activity in both normoxia and hypoxia. Conversely, LIMD1 expression downregulates HIF-1 transcriptional activity in a manner depending on PHD and 26S proteasome activities. LIMD1 family member proteins Ajuba and WTIP also bind to VHL and PHDs 1 and 3, indicating that these LIM domain-containing proteins represent a previously unrecognized group of hypoxic regulators.

AB - There are three prolyl hydroxylases (PHD1, 2 and 3) that regulate the hypoxia-inducible factors (HIFs), the master transcriptional regulators that respond to changes in intracellular O(2) tension. In high O(2) tension (normoxia) the PHDs hydroxylate two conserved proline residues on HIF-1α, which leads to binding of the von Hippel-Lindau (VHL) tumour suppressor, the recognition component of a ubiquitin-ligase complex, initiating HIF-1α ubiquitylation and degradation. However, it is not known whether PHDs and VHL act separately to exert their enzymatic activities on HIF-1α or as a multiprotein complex. Here we show that the tumour suppressor protein LIMD1 (LIM domain-containing protein) acts as a molecular scaffold, simultaneously binding the PHDs and VHL, thereby assembling a PHD-LIMD1-VHL protein complex and creating an enzymatic niche that enables efficient degradation of HIF-1α. Depletion of endogenous LIMD1 increases HIF-1α levels and transcriptional activity in both normoxia and hypoxia. Conversely, LIMD1 expression downregulates HIF-1 transcriptional activity in a manner depending on PHD and 26S proteasome activities. LIMD1 family member proteins Ajuba and WTIP also bind to VHL and PHDs 1 and 3, indicating that these LIM domain-containing proteins represent a previously unrecognized group of hypoxic regulators.

KW - Cell Hypoxia

KW - Cell Line, Tumor

KW - HEK293 Cells

KW - HeLa Cells

KW - Humans

KW - Hydroxylation

KW - Hypoxia-Inducible Factor 1, alpha Subunit/genetics

KW - Hypoxia-Inducible Factor-Proline Dioxygenases

KW - Immunoblotting

KW - Immunoprecipitation

KW - Intracellular Signaling Peptides and Proteins/genetics

KW - LIM Domain Proteins/genetics

KW - Models, Biological

KW - Polyubiquitin/metabolism

KW - Procollagen-Proline Dioxygenase/genetics

KW - Proteasome Endopeptidase Complex/metabolism

KW - Protein Binding

KW - RNA Interference

KW - Transfection

KW - Two-Hybrid System Techniques

KW - Ubiquitination

KW - Von Hippel-Lindau Tumor Suppressor Protein/genetics

U2 - 10.1038/ncb2424

DO - 10.1038/ncb2424

M3 - Article

C2 - 22286099

SN - 1465-7392

VL - 14

SP - 201

EP - 208

JO - Nature Cell Biology

JF - Nature Cell Biology

IS - 2

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