Fine-structure mapping and functional analysis of temperature-sensitive mutants in the gene encoding the herpes simplex virus type 1 immediate early protein VP175

Abstract

Herpex simplex virus (HSV)-specific proteins fall into at least 3 kinetic classes whose synthesis is sequentially and coordinately regulated. Temperature-sensitive (ts) mutants of 1 complementation group (1-2) are defective in the transition from immediate early to early and late protein synthesis. To elucidate the function of the 1-2 gene product in the HSV type 1 replicative cycle, 9 ts mutants in this group were mapped by fine-structural analysis and characterized biochemically. Physical mapping by homotypic marker rescue showed that all members of the group lie within the terminally repeated sequences of the S region of the genome. Fine-structure genetic and physical mapping permitted the mutations to be ordered within these sequences. Because the message for VP175 and the DNA template specifying this protein extend beyond the limits of the physical map of the mutations, the mutations must lie within the structural gene for VP175. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed that most members of the group overproduced the immediate early proteins VP175, -136, -110 and -63 and markedly underproduced early and late proteins at the nonpermissive temperature. In temperatrue shiftup experiments, the synthesis of early and late proteins ceased; the synthesis of immediate early proteins began again. Thus, VP175 apparently is involved in the transition from immediate early to early protein synthesis; required continuously to maintain early protein synthesis; and autoregulated, acting to inhibit immediate early protein synthesis.