EBNA-2 of herpesvirus papio diverges significantly from the type A and type B EBNA-2 proteins of Epstein-Barr virus but retains an efficient transactivation domain with a conserved hydrophobic motif

Autor: Judith J. Ryon, P D Ling, S D Hayward
Rok vydání: 1993
Předmět:
Transcriptional Activation
Herpesvirus 4
Human
viruses
Molecular Sequence Data
Immunology
Fluorescent Antibody Technique
Sequence alignment
Protein Sorting Signals
Biology
Microbiology
Conserved sequence
Structure-Activity Relationship
Transactivation
hemic and lymphatic diseases
Virology
Animals
Amino Acid Sequence
Cloning
Molecular
Promoter Regions
Genetic
Antigens
Viral
Vero Cells
Peptide sequence
Conserved Sequence
Herpesviridae
Cell Nucleus
chemistry.chemical_classification
Herpes simplex virus protein vmw65
Base Sequence
Sequence Homology
Amino Acid
Chromosome Mapping
Biological Transport
Sequence Analysis
DNA
DNA-binding domain
Molecular biology
Cell Compartmentation
Amino acid
DNA-Binding Proteins
Epstein-Barr Virus Nuclear Antigens
chemistry
Insect Science
Trans-Activators
Nuclear localization sequence
Papio
Research Article
Zdroj: Journal of Virology. 67:2990-3003
ISSN: 1098-5514
0022-538X
DOI: 10.1128/jvi.67.6.2990-3003.1993
Popis: EBNA-2 contributes to the establishment of Epstein-Barr virus (EBV) latency in B cells and to the resultant alterations in B-cell growth pattern by up-regulating expression from specific viral and cellular promoters. We have taken a comparative approach toward characterizing functional domains within EBNA-2. To this end, we have cloned and sequenced the EBNA-2 gene from the closely related baboon virus herpesvirus papio (HVP). All human EBV isolates have either a type A or type B EBNA-2 gene. However, the HVP EBNA-2 gene falls into neither the type A category nor the type B category, suggesting that the separation into these two subtypes may have been a recent evolutionary event. Comparison of the predicted amino acid sequences indicates 37% amino acid identity with EBV type A EBNA-2 and 35% amino acid identity with type B EBNA-2. To define the domains of EBNA-2 required for transcriptional activation, the DNA binding domain of the GAL4 protein was fused to overlapping segments of EBV EBNA-2. This approach identified a 40-amino-acid (40-aa) EBNA-2 activation domain located between aa 437 and 477. Transactivation ability was completely lost when the amino-terminal boundary of this domain was moved to aa 441, indicating that the motif at aa 437 to 440, Pro-Ile-Leu-Phe, contains residues critical for function. The aa 437 boundary identified in these experiments coincides precisely with a block of conserved sequences in HVP EBNA-2, and the comparable carboxy-terminal region of HVP EBNA-2 also functioned as a strong transcriptional activation domain when fused to the Gal4(1-147) protein. The EBV and HVP EBNA-2 activation domains share a mixed proline-rich, negatively charged character with a striking conservation of positionally equivalent hydrophobic residues. The importance of the individual amino acids making up the Pro-Ile-Leu-Phe motif was examined by mutagenesis. Any alteration of these residues was found to reduce transactivation efficiency, with changes at the Pro-437 and Phe-440 positions producing the most deleterious effects. Activation of the EBV latency C promoter by EBNA-2 was shown to be dependent on the presence of the carboxy-terminal activation domain. However, this requirement was generic, rather than specific, since the EBNA-2 activation domain could be replaced with those from the herpes simplex virus (HSV) VP16 protein or the EBV Rta protein. Potential karyophilic signals within EBNA-2 were examined by introducing oligonucleotides encoding positively charged amino acid groupings that might serve in this capacity into a cytoplasmic test protein, HSV delta IE175, and by examining the intracellular localization of the resulting proteins. This assay identified a strong nuclear localization signal between EBV amino acids (aa) 478 to 485, which was conserved in HVP, and a weaker noncanonical signal between EBV aa 341 to 355, which was not conserved in HVP.
Databáze: OpenAIRE
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