Antiinflammatory adaptation to hypoxia through adenosine-mediated cullin-1 deneddylation
Joseph Khoury, … , Andrew S. Neish, Sean P. Colgan
Joseph Khoury, … , Andrew S. Neish, Sean P. Colgan
Published March 1, 2007
Citation Information: J Clin Invest. 2007;117(3):703-711. https://doi.org/10.1172/JCI30049.
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Research Article Vascular biology

Antiinflammatory adaptation to hypoxia through adenosine-mediated cullin-1 deneddylation

Abstract

A major adaptive pathway for hypoxia is hypoxic preconditioning (HPC), a form of endogenous protection that renders cells tolerant to severe challenges of hypoxia. We sought to define the antiinflammatory properties of HPC. cDNA microarray analysis of lung tissue from mice subjected to hypoxia or HPC identified a cluster of NF-κB–regulated genes whose expression is attenuated by HPC. Studies using an NF-κB luciferase reporter assay confirmed a significant suppression of NF-κB activation during HPC. HPC-elicited activity was conferrable, as a soluble supernatant from HPC-treated cells, and the active fraction was purified and identified as adenosine (Ado). Guided by recent studies demonstrating bacterial inhibition of NF-κB through cullin-1 (Cul-1) deneddylation, we found a dose-dependent deneddylation of Cul-1 by Ado receptor stimulation predominantly mediated by the Ado A2B receptor subtype. Further, siRNA-mediated repression of CSN5, a subunit of the COP9 signalosome responsible for deneddylation of Cul-1, partially reversed HPC-mediated inhibition of NF-κB. Cul-1 deneddylation was evident in a murine model of HPC and lost in animals lacking extracellular Ado (Cd73–/– mice). Taken together, these results demonstrate that HPC induces extracellular accumulation of Ado and suppresses NF-κB activity through deneddylation of Cul-1. These results define a molecular regulatory pathway by which Ado provides potent antiinflammatory properties.

Authors

Joseph Khoury, Juan C. Ibla, Andrew S. Neish, Sean P. Colgan

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Figure 6

HPC deneddylates Cul-1 and stabilizes IκB in vivo in wild-type versus Cd73–/– mice.

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HPC deneddylates Cul-1 and stabilizes IκB in vivo in wild-type versus Cd...
(A) Water tissue content of whole left lungs from wild-type (left) or Cd73–/– (right) mice were analyzed, and the results confirmed the macroscopic findings shown above (Figure 5A). Data are expressed as lung water content (cc/mg tissue) from 4 mice in each group; *P < 0.01 versus normoxia, #P < 0.02 versus wild-type HPC. (B) In vivo HPC of C57BL/6 wild-type mice followed by 10 minutes severe hypoxia as described in Methods greatly deneddylates Cul-1 as compared with normoxia. CD73–/– mice do not display deneddylation of Cul-1. Of interest, mice subjected to severe hypoxia for 10 minutes without prior HPC displayed an accentuated neddylation of Cul-1, suggestive of increased NF-κB activation. Cd73–/– mice showed an increased level of neddylated to unmodified Cul-1 protein, and HPC did not lead to an increase in deneddylation. Similarly, wild-type hypoxic mice displayed decreased IκB expression and that was stabilized by PC.