We have determined the complete genomic sequence of human astrovirus serotype 1 isolated in Newcastle upon Tyne. The genome is 6813 nucleotides long and contains three sequential open reading frames (ORFs). The two closest to the 5' end are linked by a ribosomal frameshifting motif and contain sequence motifs indicative of non-structural virus proteins: serine protease and RNA-dependent RNA polymerase. A nuclear addressing sequence is also located here. The 3' ORF encodes the virion structural polypeptides as a polyprotein precursor. This genomic organization resembles that of the plant virus family Luteoviridae.

We have determined 4380 bases of the sequence from a cDNA clone containing the 3' end of feline calicivirus strain F9. We find four candidate open reading frames of which three are complete and comprise 245, 317 and 2012 nucleotides. The fourth continues toward the 5' end. We have expressed the largest complete open reading frame in E. coli. Sera raised to this antigen react specifically with the capsid protein and its intracellular precursor molecule. N-terminal sequence analysis of purified, mature capsid protein confirms this assignment and has identified the position at which precursor is cleaved.

We have used Western blotting to examine the accumulation of feline calicivirus proteins within the infected cell. Experiments using elevated growth temperature to block post-translational cleavage have demonstrated two additional high molecular weight protein bands 125 kd and 123 kd respectively which may be precursor polyproteins. Inhibition of proteolytic processing with p-fluorophenylalanine led to the accumulation of several additional protein species which may represent intermediates in the protein processing pathway. None of these proteins were related to the 62 kd major capsid protein (cP62) of the virus as judged by reaction with monoclonal antibodies. The production of a 76 kd capsid precursor protein (cpP 76) was demonstrated for the first time in FCV-infected cells. The pathway by which calicivirus polypeptides are made thus appears highly complex and may involve temporal regulation of protein synthesis as well as protein processing. Tentative identification of primary, intermediate and mature forms of virus proteins is discussed.

We report here the cloning and 3' sequence determination of feline calicivirus strain F9. Subcloning the 3' terminus enabled the production of strand specific probes for RNA synthesis. We extend the number of virus specific RNAs detected intracellularly to 8, and show that numbers 1-5 are represented as negative strands which may serve as templates in the synthesis of these RNAs.

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