Sequence relationships between Kirsten retrovirus genomes and the genomes of other murine retroviruses.

RNA sequence relationships between the genomes of the Kirsten murine sarcoma virus (MSV-K) complex, the Kirsten murine leukemia virus (MuLV-K) complex, the Gross murine leukemia virus (MuLV-G), and the Moloney murine leukemia virus (MuLV-M) were investigated. Sedimentation analyses revealed the expected 30 and 34 S RNA subunits in the MSV-K complex and a previously undetected 30 S RNA subunit accompanying the 34 S RNA subunit in the MuLV-K complex. Nucleic acid hybridization data indicated that each Kirsten virus 30 S RNA subunit had about 40% sequence homology with the RNA genome of MuLV-G, although these sequences were only partially homologous between the two 30 S subunits. In contrast, the MuLV-K 34 S RNA subunit had 96% sequence homology with the MuLV-G genome, whereas the MSV-K 34 S RNA subunit displayed only 71% sequence homology with the MuLV-G genome. Similar relationships were indicated by oligonucleotide fingerprinting. The oligonucleotide data, taken with published sequence data on the MuLV-G and MuLV-M genomes, enabled us to construct partial sequence maps of the MuLV-K 34 S RNA subunit and the MSV-K 34 and 30 S RNA subunits. The sequence arrangements indicated that (1) the MuLV-K 34 S RNA subunit is a variant of the MuLV-G genome; (2) the MSV-K 34 S RNA subunit is a recombinant molecule, which maintains the length of its leukemia virus parent; and (3) the MSV-K 30 S RNA subunit may have been generated from the MuLV-K 34 S genome by a two-stage process, culminating in the retention of parental sequences only within the U5 and U3 noncoding segments and within several amino-terminal coding segments. Further examination of published retrovirus genome sequences revealed several strategically situated sets of potential recognition signals for transcription and translation and suggested a model for genetic recombination based on mRNA splicing signals and areas of limited sequence homology. This model may explain how foreign gene elements can be inserted into retrovirus genomes to generate either functional or defective recombinant retroviruses.

Radioimmunoassay of measles virus nucleocapsid antigen.

A double antibody competitive binding radioimmunoassay (RIA) was developed as a tool for investigating the involvement of measles virus in persistent virus infections. The assay employs 125I-labelled measles virus nucleocapsids as the labelled antigen. Nucleocapsids were purified from cytoplasmic extracts of virus-infected Vero cells, treated with trypsin to prevent clumping and iodinated to a specific activity of about 15 microCi/microgram. Electrophoretic analysis of iodinated trypsin-treated nucleocapsids revealed only labelled polypeptide with a relative mol. wt. (Mr) 40 000, indicating that trypsin had cleaved the 60 000 mol. wt. native polypeptide to a 40 000 mol. wt. subunit. The assay could detect as little as 0.1 ng of nucleocapsid protein. Either native (Mr 60 000) or cleaved Mr 40 000) nucleocapsid polypeptide was detected in the assay. As much as 100 micrograms of protein from uninfected Vero cells did not react in the RIA. Another measure of specificity was the fact that 400 ng of purified nucleocapsid protein from simian virus 5, Newcastle disease virus or canine distemper virus did not react in the assay. In the RIA, nucleocapsid antigen of virus isolated from subacute sclerosing panencephalitis (SSPE) was indistinguishable from that of Edmonston strain measles virus, indicating antigenic homology between the 40 000 mol. wt. nucleocapsid polypeptide of the SSPE virus and that of measles virus.