Inhibition of ubiquitin/proteasome-dependent protein degradation by the Gly-Ala repeat domain of the Epstein-Barr virus nuclear antigen 1.
The Epstein-Barr virus (EBV) encoded nuclear antigen (EBNA) 1 is expressed in latently infected B lymphocytes that persist for life in healthy virus carriers and is the only viral protein regularly detected in all EBV associated malignancies. The Gly-Ala repeat domain of EBNA1 was shown to inhibit in cis the presentation of major histocompatibility complex (MHC) class I restricted cytotoxic T cell epitopes from EBNA4. It appears that the majority of antigens presented via the MHC I pathway are subject to ATP-dependent ubiquitination and degradation by the proteasome. We have investigated the influence of the repeat on this process by comparing the degradation of EBNA1, EBNA4, and Gly-Ala containing EBNA4 chimeras in a cell-free system. EBNA4 was efficiently degraded in an ATP/ubiquitin/proteasome-dependent fashion whereas EBNA1 was resistant to degradation. Processing of EBNA1 was restored by deletion of the Gly-Ala domain whereas insertion of Gly-Ala repeats of various lengths and in different positions prevented the degradation of EBNA4 without appreciable effect on ubiquitination. Inhibition was also achieved by insertion of a Pro-Ala coding sequence. The results suggest that the repeat may affect MHC I restricted responses by inhibiting antigen processing via the ubiquitin/proteasome pathway. The presence of regularly interspersed Ala residues appears to be important for the effect.
Levitskaya
J
J
Microbiology and Tumor Biology Center, Karolinska Institute, Box 280, S-171 77 Stockholm, Sweden.
Sharipo
A
A
Leonchiks
A
A
Ciechanover
A
A
Masucci
M G
MG
D000818:Animals; D001402:B-Lymphocytes / Q000821:virology*; D002460:Cell Line; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D019309:Epstein-Barr Virus Nuclear Antigens / Q000235:genetics / Q000378:metabolism*; D006566:Herpesviridae Infections / Q000378:metabolism* / Q000821:virology; D004854:Herpesvirus 4, Human / Q000502:physiology*; D006801:Humans; D009097:Multienzyme Complexes / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D011506:Proteins / Q000378:metabolism*; D011817:Rabbits; D012091:Repetitive Sequences, Nucleic Acid; D014412:Tumor Virus Infections / Q000378:metabolism* / Q000821:virology; D014452:Ubiquitins / Q000378:metabolism*
D019309:Epstein-Barr Virus Nuclear Antigens; D009097:Multienzyme Complexes; D011506:Proteins; D014452:Ubiquitins; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex