Proteasomal degradation of human CYP1B1: effect of the Asn453Ser polymorphism on the post-translational regulation of CYP1B1 expression

Mol Pharmacol. 2005 Feb;67(2):435-43. doi: 10.1124/mol.104.006056. Epub 2004 Oct 14.

Abstract

Allelic variations in CYP1B1 are reported to modulate the incidence of several types of cancer. To provide a mechanistic basis for this association, we investigated the impact of nonsilent allelic changes on the intracellular levels and post-translational regulation of CYP1B1 protein. When transiently expressed in COS-1 cells, either in the presence or absence of recombinant cytochrome P450 reductase, the cellular level of the CYP1B1.4 allelic variant (containing a Ser at the amino acid position 453; Ser453) was 2-fold lower compared with the other four allelic CYP1B1 proteins (containing Asn453), as analyzed by both immunoblotting and ethoxyresorufin O-deethylase activity. This difference was caused by post-translational regulation; as in the presence of cycloheximide, the rate of degradation of immunodetectable and enzymatically active CYP1B1.4 was distinctly faster than that of CYP1B1.1. Pulse-chase analysis revealed that the half-life of CYP1B1.4 was a mere 1.6 h compared with 4.8 h for CYP1B1.1. The presence of the proteasome inhibitor MG132 [N-benzoyloxycarbonyl (Z)-Leu-Leuleucinal] increased the stability not only of immunodetectable CYP1B1, but also--unexpectedly given the size of the proteasome access channel--increased the stability of enzymatically active CYP1B1. The data presented herein also demonstrate that CYP1B1 is targeted for its polymorphism-dependent degradation by polyubiquitination but not phosphorylation. Our results importantly provide a mechanism to explain the recently reported lower incidence of endometrial cancer in individuals carrying the CYP1B1*4 compared with the CYP1B1*1 haplo-type. In addition, the mechanistic paradigms revealed herein may explain the strong overexpression of CYP1B1 in tumors compared with nondiseased tissues.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles
  • Amino Acid Substitution / genetics
  • Animals
  • Aryl Hydrocarbon Hydroxylases
  • Asparagine / genetics*
  • COS Cells
  • Chlorocebus aethiops
  • Cytochrome P-450 CYP1B1
  • Cytochrome P-450 Enzyme System / biosynthesis
  • Cytochrome P-450 Enzyme System / genetics*
  • Cytochrome P-450 Enzyme System / metabolism*
  • Enzyme Stability / genetics
  • Gene Expression Regulation, Enzymologic / genetics
  • Polymorphism, Genetic / genetics*
  • Proteasome Endopeptidase Complex / genetics
  • Proteasome Endopeptidase Complex / metabolism*
  • Protein Processing, Post-Translational / genetics*
  • Serine / genetics*

Substances

  • Serine
  • Asparagine
  • Cytochrome P-450 Enzyme System
  • Aryl Hydrocarbon Hydroxylases
  • CYP1B1 protein, human
  • Cytochrome P-450 CYP1B1
  • Proteasome Endopeptidase Complex