Peptides inhibitory for the transcriptional regulatory function of human papillomavirus E2.
OBJECTIVE
Human papillomavirus (HPV) infections are associated with cervical neoplasia. Cellular and viral proteins are known to interact with the papillomavirus E2 protein to initiate transcription and DNA replication in the HPV life cycle. Our aim was to identify peptides that bind to the HPV16 E2 protein and thereby inhibit its ability to alter the transcriptional activity of other genes.


METHODS
The HPV16 E2 protein was expressed and purified to near homogeneity in bacteria. We screened a phage display library of random peptides for ones that bound to HPV16 E2 protein. Among the isolated phage clones, we found that tryptophan-rich peptide sequences appeared repetitively in successive cycles of phage library panning. Replacement of the tryptophan amino acids in these dodecapeptides reduced the degree to which these peptides bound to the E2 protein. These E2-binding peptides were tested for their ability to inhibit the transcriptional regulatory function of E2 in a test cell line, which contained an E2 gene and a luciferase reporter gene driven by an E2-dependent transcriptional promoter.


RESULTS
Delivery of four of the E2 binding peptides into the intracellular compartment of the test cell line resulted in suppression of the E2-dependent luciferase expression. Deletion of the tryptophan residues from these peptides reduced their E2 binding and their ability to suppress E2-dependent luciferase expression in the test cell line.


CONCLUSIONS
These results suggest a strategy for the development of chemical inhibitors of E2-dependent transcription of viral genes in HPV-infected cells as an approach to the therapy of chronic HPV infections.
Fujii
Takuma
T
Yale University, New Haven, Connecticut 06520, USA.
Austin
David
D
Guo
David
D
Srimatkandada
Srinivasan
S
Wang
Tao
T
Kubushiro
Kaneyuki
K
Masumoto
Nobuo
N
Tsukazaki
Katsumi
K
Nozawa
Shiro
S
Deisseroth
Albert B
AB
D002478:Cells, Cultured; D004261:DNA Replication / Q000187:drug effects; D004279:DNA, Viral / Q000037:antagonists & inhibitors / Q000378:metabolism; D004268:DNA-Binding Proteins; D015967:Gene Expression Regulation, Viral; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000378:metabolism*; D027383:Papillomaviridae / Q000187:drug effects / Q000502:physiology; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology*; D019151:Peptide Library; D014158:Transcription, Genetic / Q000187:drug effects*; D014364:Tryptophan / Q000737:chemistry / Q000378:metabolism; D014764:Viral Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D014779:Virus Replication / Q000187:drug effects
D004279:DNA, Viral; D004268:DNA-Binding Proteins; C054855:E2 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D010446:Peptide Fragments; D019151:Peptide Library; D014764:Viral Proteins; D014364:Tryptophan; D008156:Luciferases