Isolation and characterization of temperature-sensitive mutants of adenovirus type2

Abstract

Temperature-sensitive mutants (14) of human adenovirus type 2, which differed in their plaquing efficiencies at the permissive and nonpermissive temperatures by 4-5 orders of magnitude, were isolated. These mutants, which could be assigned to 7 complementation groups, were tested for their capacity to synthesize adenovirus DNA at the nonpermissive temperature. Three mutants in 3 different complementation groups were deficient in viral DNA synthesis. The DNA-negative mutant H2ts206 complemented the DNA-negative mutants H5ts36 and H5ts125, whereas mutant H2ts201 complemented H5ts36 only. Among the DNA-negative mutants, H2ts206 synthesized the smallest amount of viral DNA at the nonpermissive temperature (39.5.degree. C). Data obtained in temperature shift experiments indicated that a very early function was involved in temperature sensitivity. In keeping with this observation, early virus-specific mRNA was not detected in cells [HeLa, KB human oral carcinoma] infected with H2ts206 and maintained at 39.5.degree. C. Prolonged (52 h) incubation of cells infected with H2ts206 at the nonpermissive temperature led to the synthesis of a high-MW form of viral DNA.