We have examined whether sympathetic neurones that have lost the potential to be rescued by protein and RNA synthesis inhibitors after a period of nerve growth factor (NGF) deprivation are irreversibly committed to die. We found that 15 h after withdrawal of NGF from 7-day cultures of neonatal rat superior cervical ganglion neurones, 50% of the neurones lost the potential to be rescued by cycloheximide but that NGF rescued most of the neurones. By 22 h after NGF withdrawal, only 10% of the neurones were rescued by inhibition of macromolecular synthesis with cycloheximide, puromycin, or actinomycin D, but as many as 60-80% of the neurones were rescued by NGF. This is after the time at which a DNA "ladder," consistent with cell death by apoptosis, was first detected (18 h). As long as 27 h of NGF withdrawal was required before 50% of the neurones lost the potential to be rescued by NGF. The survival-promoting agent 8-(4-chlorophenylthio)cyclic AMP (CPTcAMP) or depolarization with 50 mM KCl (HK) rescued neurones with kinetics similar to those of NGF, and rescue by all three agents did not require protein synthesis. Thus, NGF, CPTcAMP, and HK can rescue neurones deprived of NGF at much later times than either protein or RNA synthesis inhibitors by acting at the posttranslational level, a finding suggesting that initiation of the cell death programme in sympathetic neurones is not an irreversible step.

SUMMARYPurified DNAs from Marek's disease virus (MDV) and the herpesvirus of turkeys (HVT) were randomly sheared and cloned into the M 13 bacteriophage. Two-hundred and ten MDV and 130 HVT clones were sequenced to give representative samples of the genome sequences. The predicted amino acid sequences from these gammaherpesviruses were compared to known sequences from other herpesviruses using computer analysis. Thirty-five MDV and 24 HVT genes were identified by comparison with varicella-zoster virus (VZV), an alphaherpesvirus. However, only 14 MDV and seven HVT genes, giving generally lower homology scores, were found by comparison with Epstein-Barr virus (EBV), a gammaherpesvirus, indicating that MDV and HVT sequences bear greater similarity to VZV than to EBV sequences. A number of sequences were mapped by hybridizing labelled M13 clones to Southern blots of restriction fragments of MDV or HVT DNA. The results were consistent with the MDV and HVT genomes being collinear with VZV.

Marek's disease herpesvirus (MDV) can cause either a productive-restrictive or lytic infection, a latent infection or can transform thymus-derived lymphocytes. RNA was extracted from infected chicken embryo fibroblasts (CEF) or from lymphoblastoid tumour cell lines. Some of the infected CEF were treated with 200 micrograms/ml cycloheximide to identify immediate early (IE) transcripts, and others with 1 microM 1-(2-fluoro-2-deoxy-B-D-arabinofuranosyl)-5-methyluracil (FMAU), an inhibitor of herpesvirus DNA synthesis to identify early transcripts. An extensive Northern blot analysis was carried out using DNA probes spanning almost the complete MDV genome. In the lytically infected CEF at least 66 discrete transcripts were detected, ranging in size from 9.1 kb to 0.6 kb. Eleven IE transcripts were identified, of which 8 were mapped in the genome segment consisting of the IRL, IRS, US and TRS. Six transcripts were identified as early genes. In the MD lymphoblastoid cell lines MDCC-HPI, a non-producer cell line, and MDCC-CU41, a non-expression cell line, 4 and 7 transcripts were detected, respectively. These RNAs were transcribed from IE genes located mainly in the repeat sequences flanking UL and US and in US. Treatment of the lymphoblastoid cell lines with 20 micrograms/ml 5-iodo-2-deoxyuridine resulted in the additional transcription of 1 RNA species in HPI and 9 in CU41. Most of the transcripts present in lytically infected cells were also detected in MDCC-CU36, a cell line with a high percentage of antigen-positive cells (expression cell line).

SUMMARYThe production and properties of three monoclonal antibodies, designated LP 1, LP4 and AP2, directed against non-glycosylated polypeptides of herpes simplex virus type 2 are described. LP1 is specific for polypeptide VP16 and cross-reacts with HSV-1 ; LP4 reacts with the major DNA-binding protein and is type-specific. AP2 is directed against the major capsid antigen of HSV-1 and HSV-2.

Evidence is presented showing that the 92,000-dalton glycoprotein (g92K) induced by herpes simplex virus (HSV) type 2 has properties distinct from those assigned to any other HSV glycoprotein. First, the carbohydrate composition and extent of sulfation differ from those of glycoproteins D and E. Second, two clonally unrelated monoclonal antibodies, AP1 and LP5, shown in this paper to specifically immunoprecipitate g92K, do not react with any of the known processed forms of glycoproteins B, C, D, and E. Third, by using HSV type 1/HSV type 2 intertypic recombinants and a simple radioimmunoassay, the target antigen of the two monoclonal antibodies was shown to map in the same region as g92K (0.846 to 0.924). Fourth, the intertypic recombinant R12-3 was shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of infected cells to induce the HSV type 2 g92K and HSV type 1 gD and gE, whereas R12-1, which did not induce g92K, induced HSV-2 gE and an altered gD, providing genetic evidence that g92K is encoded, at least in part, by a different region of the genome from that encoding gD and gE.