Inhibition of the Catalytic Activity of Hypoxia-Inducible Factor-1α-Prolyl-Hydroxylase 2 by a MYND-Type Zinc Finger

  1. Kyung-Ok Choi,
  2. Taekyong Lee,
  3. Naery Lee1,
  4. Ji-Hyun Kim,
  5. Eun Gyeong Yang,
  6. Jung Min Yoon,
  7. Jin Hwan Kim,
  8. Tae Gyu Lee and
  9. Hyunsung Park
  1. Department of Life Science, University of Seoul, Seoul, Korea (K.-O.C., T.L., N.L., H.P.); Life Science Division, Korea Institute of Science and Technology, Seoul, Korea (J.-H.K., E.G.Y.); and the Division of Drug Discovery, CrystalGenomics, Inc., Daejeon City, Korea (J.M.Y., J.H.K., T.G.L.)
  1. Address correspondence to:
    Dr. Hyunsung Park, Department of Life Science, University of Seoul, 90 Cheonnong-dong, Tongdaemun-gu, Seoul 130-743, Korea. E-mail: hspark{at}uos.ac.kr.

Abstract

Hypoxia-induced gene expression is initiated when the hypoxia-inducible factor-1 (HIF-1) α subunit is stabilized in response to a lack of oxygen. An HIF-1α-specific prolyl-hydroxylase (PHD) catalyzes hydroxylation of the proline-564 and/or -402 residues of HIF-1α by an oxygen molecule. The hydroxyproline then interacts with the ubiquitin E3 ligase von Hippel Lindau protein and is degraded by an ubiquitin-dependent proteasome. PHD2 is the most active of three PHD isoforms in hydroxylating HIF-1α. Structural analysis showed that the N-terminal region of PHD2 contains a Myeloid translocation protein 8, Nervy, and DEAF1 (MYND)-type zinc finger domain, whereas the catalytic domain is located in its C-terminal region. We found that deletion of the MYND domain increased the activity of both recombinant PHD2 protein and in vitro-translated PHD2. The zinc chelator N,N,N′,N′-tetrakis(2-pyridylmethyl)ethylenediamine augmented the activity of wild-type PHD2-F but not that of PHD2 lacking the MYND domain, confirming that the zinc finger domain is inhibitory. Overexpression of PHD2 lacking the MYND domain caused a greater reduction in the stability and function of HIF-1α than did overexpression of wild-type PHD2, indicating that the MYND domain also inhibits the catalytic activity of PHD2 in vivo.

Footnotes

  • This work was supported by grant KRF-2002-041-C00207 from the Korean Research Foundation (to H.P.).

  • K.-O.C., T.L., and N.L. contributed equally to this study.

  • Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.

  • doi:10.1124/mol.105.015271.

  • ABBREVIATIONS: HIF-1, hypoxia-inducible factor-1; ODD, oxygen dependent degradation domain; VHL, von Hippel Lindau; MYND, Myeloid translocation protein 8, Nervy, and DEAF1; DMEM, Dulbecco's modified Eagle's medium; TPEN, N,N,N′,N′-tetrakis(2-pyridylmethyl)ethylenediamine; GST, glutathione S-transferase; PAGE, polyacrylamide gel electrophoresis; MALDI-TOF, matrix-assisted laser desorption ionization/time of flight.

  • ↵1 Current affiliation: Biologics Evaluation Department, Biologics Standardization Division, Korea Food and Drug Administration, Seoul, Korea.

    • Received May 26, 2005.
    • Accepted September 9, 2005.
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  1. Published online before print September 9, 2005, doi: 10.1124/mol.105.015271 Molecular Pharmacology December 2005 vol. 68 no. 6 1803-1809
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