Control of Protein Synthesis in Herpesvirus-infected Cells: Analysis of the Polypeptides Induced by Wild Type and Sixteen Temperature-sensitive Mutants of HSV strain 17

Abstract

SUMMARYThe polypeptides induced in cells infected with a Glasgow isolate of HSV-I (I 7 syn +) have been characterized by SDS polyacrylamide gel electrophoresis.Study of the kinetics of synthesis in three cell lines has detected a total of 52 polypeptides, 33 of which can be identified in polypeptide profiles of purified virions. These include six low mol. wt. polypeptides that have not been previously reported. Several polypeptides were labelled with glucosamine in infected BHK cells.The different polypeptide patterns obtained at permissive (31 °C) and nonpermissive (38 °C) temperature in cells infected with 16 temperature-sensitive (ts) mutants are reported. The effect of multiplicity of infection (m.o.i.) on the polypeptide profile has been examined for two of the DNA -ve mutants: below ten, the profile varied with the m.o.i, whereas above ten it was constant. All mutants were therefore examined at an m.o.i, of approx. 20. Mutants from the same complementation group showed very similar profiles.A number of general conclusions concerning control of protein synthesis in HSV infected cells can be made: (I) As most of the 16 ts mutants affected the synthesis of several or many polypeptides it follows that a large proportion of the genome specifies controlling functions. (2) The high frequency with which some polypeptides were affected suggests they are at or near the terminus of biosynthetic pathways which are under multiple control. (3) Conversely, some polypeptides were affected with a low frequency suggesting that their synthesis is not dependent on the expression of many virus functions. (4) Several individual ts mutations lead to the synthesis of increased amounts of different large polypeptides. (5) Analysis of every band detectably affected by at least one ts mutation has disclosed nine classes of dependence relationship between polypeptide synthesis and the DNA phenotype of the mutants, illustrating that this relationship is complex and different for different polypeptides. (6) The inhibition of host protein synthesis by the virus may not be a simple single step process.

Characterization of the IE110 Gene of Herpes Simplex Virus Type 1

“…This is considerably lower than the previous estimate of 110000 obtained from gel electrophoretic mobility (Marsden et al, 1976). However, such revisions of molecular weights have occurred frequently in HSV sequence analysis (see, for example, McGeoch et al, 1985).…”

“…Conditions for increased production of IE polypeptides using cycloheximide block have previously been described (Preston, 1979). Immunoprecipitations were carried out as described (Frame et al, 1986) and analysed on 5 to 12.5 ~ gradient SDS-polyacrylamide gels (Marsden et al, 1976).…”

One Functional Copy of the Long Terminal Repeat Gene Specifying the Immediate-early Polypeptide IE 110 Suffices for a Productive Infection of Human Foetal Lung Cells by Herpes Simplex Virus

“…After 10 min incubation the nuclei were pelleted by centrifugation at 2000 rev/min for 5 min and the supernatant discarded. The nuclei were resuspended and boiled in denaturing buffer (Marsden et al, 1976).…”

Identification of Two Herpes Simplex Virus Type 1-induced Proteins (21K and 22K) which Interact Specifically with the a Sequence of Herpes Simplex Virus DNA

“…T h e only sequences w h i c h p G 3 b , p R 9 a n d p R 1 0 share are those c o n t a i n e d w i t h i n the 400 b p a sequence. Previous kinetic studies s h o w e d t h a t late in infection, after the onset o f H S V D N A synthesis, a 2 1 K p o l y p e p t i d e is i n d u c e d in large a m o u n t s a n d a 22K p o l y p e p t i d e in lesser a m o u n t s (Marsden et al, 1976). We therefore tested whether the 21K and 22K p01ypeptides which bind specifically to the a sequence were made early or late in infection.…”

“…Kinetic studies of HSV polypeptides induced in infected cells (Marsden et al, 1976) established that they are induced only late in infection. The observation (Fig.…”