Degradation of the tumor suppressor protein p53 by the ubiquitin-mediated proteolytic system requires a novel species of ubiquitin-carrier protein, E2.
The tumor suppressor protein p53 is extremely unstable in most cell lines. In contrast, many mutant and oncogenic species of the protein are stable. The degradation of p53 in vivo requires metabolic energy; however, the proteolytic system(s) involved have not been identified. The ubiquitin system has been implicated in the degradation of p53 in vitro. The degradation is stimulated significantly by the human papillomavirus (HPV) oncoprotein E6 that associates with p53 and facilitates conjugate formation and subsequent degradation. Complex formation between E6 and p53 is promoted by a cellular protein designated E6-associated protein (E6-AP). Initial dissection of the conjugation process have demonstrated a role for the ubiquitin-activating enzyme, E1, but the ubiquitin-carrier protein (E2, UBC) and the ubiquitin protein ligase, E3, have not been identified. In this study, we report that a novel species of ubiquitin-carrier protein designated E2-F1 (Blumenfeld, N., Gonen, H., Mayer, A., Smith, C., Siegel, N.R., Schwartz, A.L., and Ciechanover, A. (1994) J. Biol. Chem. 269, 9574-9581) is involved in the conjugation and degradation of p53. This E2 enzyme recognizes non-"N-end rule" protein substrates and appears to mediate their conjugation via a novel species of E3. The process of recognition appears to be selective; E2-F1 is not required for the conjugation and degradation of human N-myc. The involvement of E2-F1 in the in vitro process appears to be physiologically meaningful and to reproduce the in vivo process; mutant species of p53 that do not interact with E6 and are stable in vivo are not recognized by the cell free system.
Ciechanover
A
A
Department of Biochemistry, Faculty of Medicine, Technion-Israel Institute of Technology, Haifa.
Shkedy
D
D
Oren
M
M
Bercovich
B
B
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D003001:Cloning, Molecular; D006801:Humans; D008025:Ligases / Q000302:isolation & purification / Q000378:metabolism*; D051379:Mice; D016271:Proto-Oncogene Proteins c-myc / Q000302:isolation & purification / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D012156:Reticulocytes / Q000201:enzymology; D013379:Substrate Specificity; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes; D014452:Ubiquitins / Q000378:metabolism*
D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D011994:Recombinant Proteins; D016159:Tumor Suppressor Protein p53; D014452:Ubiquitins; D000255:Adenosine Triphosphate; D044763:Ubiquitin-Conjugating Enzymes; D008025:Ligases