Chaperone-dependent E3 ligase CHIP ubiquitinates and mediates proteasomal degradation of soluble guanylyl cyclase

Tian Xia, Christiana Dimitropoulou, Jingmin Zeng, Galina N. Antonova, Connie Snead, Richard C. Venema, David Fulton, Shuibing Qian, Cam Patterson, Andreas Papapetropoulos, John D. Catravas

Research output: Contribution to journalArticle

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Abstract

The nitric oxide receptor soluble guanylyl cyclase (sGC) exists in multimeric protein complexes, including heat shock protein (HSP) 90 and endothelial nitric oxide synthase. Inhibition of HSP90 by geldanamycin causes proteasomal degradation of sGC protein. In this study, we have investigated whether COOH terminus of heat shock protein 70-interacting protein (CHIP), a co-chaperone molecule that is involved in protein folding but is also a chaperone-dependent ubiquitin E3 ligase, could play a role in the process of degradation of sGC. Transient overexpression of CHIP in COS-7 cells degraded heterologous sGC in a concentration-related manner; this downregulation of sGC was abrogated by the proteasome inhibitor MG-132. Transfection of tetratricopeptide repeats and U-box domain CHIP mutants attenuated sGC degradation, suggesting that both domains are indispensable for CHIP function. Results from immunoprecipitation and indirect immunofluorescent microscopy experiments demonstrated that CHIP is associated with sGC, HSP90, and HSP70 in COS-7 cells. Furthermore, CHIP increased the association of HSP70 with sGC. In in vitro ubiquitination assays using purified proteins and ubiquitin enzymes, E3 ligase CHIP directly ubiquitinated sGC; this ubiquitination was potentiated by geldanamycin in COS-7 cells, followed by proteasomal degradation. In rat aortic smooth muscle cells, endogenous sGC was also degraded by adenovirus-infected wild-type CHIP but not by the chaperone interaction-deficient K30A CHIP, whereas CHIP, but not K30A, attenuated sGC expression in, and nitric oxide donor-induced relaxation of, rat aortic rings, suggesting that CHIP plays a regulatory role under physiological conditions. This study reveals a new mechanism for the regulation of sGC, an important mediator of cellular and vascular function.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume293
Issue number5
DOIs
StatePublished - Nov 1 2007
Externally publishedYes

Fingerprint

Ubiquitin-Protein Ligases
COS Cells
Ubiquitination
Soluble Guanylyl Cyclase
HSP90 Heat-Shock Proteins
Proteins
Proteasome Inhibitors
HSP70 Heat-Shock Proteins
Nitric Oxide Donors
Nitric Oxide Synthase Type III
Protein Folding
Immunoprecipitation
Adenoviridae
Smooth Muscle Myocytes
Transfection
Blood Vessels
Microscopy
Down-Regulation

Keywords

  • COOH terminus of heat shock protein 70-interacting protein

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)

Cite this

Chaperone-dependent E3 ligase CHIP ubiquitinates and mediates proteasomal degradation of soluble guanylyl cyclase. / Xia, Tian; Dimitropoulou, Christiana; Zeng, Jingmin; Antonova, Galina N.; Snead, Connie; Venema, Richard C.; Fulton, David; Qian, Shuibing; Patterson, Cam; Papapetropoulos, Andreas; Catravas, John D.

In: American Journal of Physiology - Heart and Circulatory Physiology, Vol. 293, No. 5, 01.11.2007.

Research output: Contribution to journalArticle

Xia, Tian ; Dimitropoulou, Christiana ; Zeng, Jingmin ; Antonova, Galina N. ; Snead, Connie ; Venema, Richard C. ; Fulton, David ; Qian, Shuibing ; Patterson, Cam ; Papapetropoulos, Andreas ; Catravas, John D. / Chaperone-dependent E3 ligase CHIP ubiquitinates and mediates proteasomal degradation of soluble guanylyl cyclase. In: American Journal of Physiology - Heart and Circulatory Physiology. 2007 ; Vol. 293, No. 5.
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AU - Snead, Connie

AU - Venema, Richard C.

AU - Fulton, David

AU - Qian, Shuibing

AU - Patterson, Cam

AU - Papapetropoulos, Andreas

AU - Catravas, John D.

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