Characterization of DNA sequence-common and sequence-specific proteins binding to cis-acting sites for cleavage of the terminal a sequence of the herpes simplex virus 1 genome.
The terminal 500-base-pair alpha sequence of the herpes simplex virus 1 genome contains signals for cleavage (Pac1 and Pac2) of unit-length DNA molecules from concatemers in unique stretches of sequences designated Ub and Uc, respectively, and a cis site for cleavage designated DR1. We report that nuclear extracts from infected cells contain factors which form two DNA-virus-specific protein complexes with components of the a sequence. Purification of the factors forming the V2 complex yielded a protein with an apparent molecular weight of 82,000 binding to DNA in a non-sequence-specific manner. Addition of Mg2+ to the purified protein-DNA probe mixture resulted in exonucleolytic degradation of the DNA. The protein was identified as the virus-specific DNase with monoclonal antibody specific for the viral enzyme. The purification of the proteins forming the V4 complex yielded two proteins with molecular weights of greater than 250,000 and 140,000 corresponding to infected cell protein 1 and to an as yet unidentified protein, respectively. These proteins formed two DNA sequence-common bands with a number of DNA probes and one sequence-specific band with probes containing both Pac2 and DR1 but not with probes containing either site alone or Pac1 and DR1. Since the DNA probe containing Pac2 and DR1 inserted into viral genome or into amplicons induced specific cleavage of the DR1 sequence whereas the nonreactive probes failed to induce the cleavage, the formation of this sequence-specific DNA-protein complex is significant and may reflect a DNA-protein interaction essential for cleavage. The possible role of the proteins identified in this study for the cleavage-packaging of viral DNA into capsids is presented.
Chou
J
J
Marjorie B. Kovler Viral Oncology Laboratories, University of Chicago, Illinois 60637.
Roizman
B
B
D015342:DNA Probes; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D006367:HeLa Cells; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D012045:Regulatory Sequences, Nucleic Acid; D018139:Simplexvirus / Q000235:genetics* / Q000254:growth & development; D014764:Viral Proteins / Q000302:isolation & purification / Q000378:metabolism*; D014779:Virus Replication
D015342:DNA Probes; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D014764:Viral Proteins