Adenovirus DNA-binding protein in cells infected with wild-type 5 adenovirus and two DNA-minus, temperature-sensitive mutants, H5ts125 and H5ts149

Abstract

H5ts125, an adenovirus type 5 conditionally lethal, temperature-sensitive (ts) mutant defective in initiation of DNA synthesis, was further characterized and the single-strand-specific DNA-binding (72,000 MW) protein was studied as to whether it is coded by the mutated viral gene. When H5ts125-infected [human oral carcinoma KB] cells were labeled with [35S]methionine at 32.degree. C and then incubated without isotope at 39.5.degree. C, the mutant''s nonpermissive temperature, the 72,000 MW polypeptide was progressively degraded. Immunofluorescence examination of cells infected with wild-type virus, H5ts125 and H5ts149 (a 2nd, unique DNA-minus mutant) showed that immunologically reactive DNA-binding protein was barely detectable in H5ts125-infected cells at 39.5.degree. C but was present in wild-type- and H5ts149-infected cells. This examination also revealed that the protein made at 32.degree. C in H5ts125-infected cells lost its ability to bind specific DNA-binding protein antibody when the infected cells were shifted to 39.5.degree. C, and that if H5ts125-infected cells were shifted from the restrictive temperature to 32.degree. C even in the presence of cycloheximide to stop protein synthesis, immunologically reactive DNA-binding protein reappeared.