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Our previous data suggested that translation in an EMC virus RNA-programmed cell-free system from Krebs-2 cells is initiated predominantly at a single site and that the earliest amino acid sequences synthesized correspond to non-structural 'leader' polypeptides p14 and p12 [(1982) FEBS Lett. 141, 153-1561. Here, polypeptides p14 and p12 were labelled in vitro by tritiated amino acids, isolated and subjected to automated Edman degradation. Both polypeptides (after the loss of the N-terminal methionine) were shown to contain alanine in position 1 and glutamic acid in positions 5 and 7. These and other data demonstrate that p14 and p12 share a common N-terminal sequence. This sequence coincides precisely with the N-terminus of EMC virus polyprotein sequence deduced from the primary structure of the viral genome [(1984) Nucleic Acids Res., in press]. Thus, the single initiation site operating in our translation system corresponds to the start of the polyprotein molecule. Picornavirus Cell-free protein synthesis Translation initiation Radiochemical sequence analysisPolyprotein processing

Summary. Hepatitis E virus (RNA and antigen) was detected in serial passagesof FRhK-4 cells after they had been co-cultivated with primary kidney cells derived from cynomolgus monkeys experimentally infected with this virus.Hepatitis E virus (HEV) is the etiological agent of enterically transmitted human hepatitis E, acute jaundice disease widely spread in many tropical countries [2,4]. Under laboratory conditions the infection has been reproduced in several species of non-human primates after inoculation with HEV-containing clinical specimens [1,5,8,11]. Numerous attempts to grow this virus in cultivated cells using conventional cell culture inoculation techniques have so far proved unsuccessful. In this paper we report the establishment of cell lines which harbour the HEV.The experiments were performed as follows: (1) Cynomolgus monkeys (Macaca faseicularis) were inoculated with the HEV (2598 Osh strain originated from Soviet Central Asia) as described earlier [1,3]. (2) As soon as the levels of serum alanine aminotransferase became pathologically elevated the animals were sacrified; the kidneys were removed aseptically and used for preparation of primary cell monolayers following the usual procedure [6]. 'The cells were cultivated for 14 days at 37 °C in 500ml flat glass flasks in 0.5% lactalbumin hydrolysate medium supplemented with 15% of fetal calf serum (Flow). (3) The primary cells were sub-passaged 1-2 times in the Eagle's MEM with 10% of fetal calf serum at 7-8 day intervals. (4) These cells were allowed to proliferate until sub-confluent monolayers had been formed, then the medium was decanted and a suspension of fresh fetal rhesus monkey kidney (FRhK-4) cells [13]