Degradation of the E7 human papillomavirus oncoprotein by the ubiquitin-proteasome system: targeting via ubiquitination of the N-terminal residue.
The E7 oncoprotein of the high risk human papillomavirus type 16 (HPV-16), which is etiologically associated with uterine cervical cancer, is a potent immortalizing and transforming agent. It probably exerts its oncogenic functions by interacting and altering the normal activity of cell cycle control proteins such as p21WAF1, p27KIP1 and pRb, transcriptional activators such as TBP and AP-1, and metabolic regulators such as M2-pyruvate kinase (M2-PK). Here we show that E7 is a short-lived protein and its degradation both in vitro and in vivo is mediated by the ubiquitin-proteasome pathway. Interestingly, ubiquitin does not attach to any of the two internal Lysine residues of E7. Substitution of these residues with Arg does not affect the ability of the protein to be conjugated and degraded; in contrast, addition of a Myc tag to the N-terminal but not to the C-terminal residue, stabilizes the protein. Also, deletion of the first 11 amino acid residues stabilizes the protein in cells. Taken together, these findings strongly suggest that, like MyoD and the Epstein Barr Virus (EBV) transforming Latent Membrane Protein 1 (LMPI), the first ubiquitin moiety is attached linearly to the free N-terminal residue of E7. Additional ubiquitin moieties are then attached to an internal Lys residue of the previously conjugated molecule. The involvement of E7 in many diverse and apparently unrelated processes requires tight regulation of its function and cellular level, which is controlled in this case by ubiquitin-mediated proteolysis.
Reinstein
E
E
Department of Biochemistry and the Rappaport Family Institute for Research in the Medical Sciences, The Bruce Rappaport Faculty of Medicine, Haifa, Israel.
Scheffner
M
M
Oren
M
M
Ciechanover
A
A
Schwartz
A
A
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D002474:Cell-Free System; D002522:Chlorocebus aethiops; D006367:HeLa Cells; D006801:Humans; D008025:Ligases / Q000378:metabolism; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D009097:Multienzyme Complexes / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D050725:Papillomavirus E7 Proteins; D017434:Protein Structure, Tertiary; D011817:Rabbits; D044763:Ubiquitin-Conjugating Enzymes; D014452:Ubiquitins / Q000235:genetics / Q000378:metabolism*
D009097:Multienzyme Complexes; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D014452:Ubiquitins; C059731:oncogene protein E7, Human papillomavirus type 16; C046288:ubiquitin carrier proteins; D000255:Adenosine Triphosphate; D044763:Ubiquitin-Conjugating Enzymes; D008025:Ligases; D008239:Lysine