Spleen focus-forming Friend virus: identification of genomic RNA and its relationship to helper virus RNA.

The genome of the defective, murine spleen focus-forming Friend virus (SFFV) was identified as a 50S RNA complex consisting of 32S RNA monomers. Electrophoretic mobility and the molecular weights of unique RNase T1-resistant oligonucleotides (T1-oligonucleotides) indicated that the 32S RNA had a complexity of about 7.4 kilobases. Hybridization with DNA complementary to Friend murine leukemia virus (Fr-MLV) has distinguished two sets of nucleotide sequences in 32S SFFV RNA, 74% which were Fr-MLV related and 26% which were SFFV specific. By the same method, SFFV RNA was 48% related to Moloney MLV. We have resolved 23 large T1-oligonucleotides of SFFV RNA and 43 of Fr-MLV RNA. On the basis of the relationship between SFFV and Fr-MLV RNAs, the 23 SFFV oligonucleotides fell into four classes: (i) seven which had homologous equivalents in Fr-MLV RNA; (ii) six more which could be isolated from SFFV RNA-Fr-MLV cDNA hybrids treated with RNases A and T1; (iii) eight more which were isolated from hybrids treated with RNases A and T1; and (iv) two which did not have Fr-MLV-related counterparts. Surprisingly, the two class iv oligonucleotides had homologous counterparts in the RNA of six amphotropic MLV's including mink cell focus-forming and HIX-MLVs analyzed previously. The map locations of the 23 SFFV T1-oligonucleotides relative to the 3' polyadenylic acid coordinate of SFFV RNA were deduced from the size of the smallest polyadenylic acid-tagged RNA fragment from which a given oligonucleotide was isolated. The resulting oligonucleotide map could be divided roughly into three segments: two terminal segments which are mosaics of oligonucleotides of classes i, ii, and iii and an internal segment between 2 and 2.5 kilobases from the 3' end containing the two oligonucleotides shared with amphotropic MLVs. Since SFFV RNA consists predominantly of sequence elements related to ecotropic and amphotropic helper-independent MLVs, it would appear that the transforming gene of SFFV is not a major specific sequence unrelated to genes of helper viruses, as is the case with Rous sarcoma and probably withe other defective sarcoma and acute leukemia viruses.

Analysis of translational products of Friend strain of spleen focus-forming virus.

We have analyzed normal rat kidney cells nonproductively infected with the Friend strain of spleen focus-forming virus (SFFV) for the presence of murine leukemia virus-specific type C viral proteins. SFFV was found to code for the p15 and p12 proteins of Friend-murine leukemia virus as determined by immunological typing of their antigenic determinants. Molecular-size analysis of p15 and p12 proteins in SFFV nonproductively infected normal rat kidney cells indicated that these proteins are translated as parts of polyprotein molecules. The apparent molecular weights of the polypeptides bearing p12 antigenic determinants revealed the presence of translational products of the SFFV genome that could not be accounted for by know type C virus helper structural proteins.

Helper-independent mink cell focus-inducing strains of Friend murine type-C virus: potential relationship to the origin of replication-defective spleen focus-forming virus.

Recent studies have indicated that both the replication-defective spleen focus-forming virus (SFFV) in the Friend virus complex and the helper-independent mink cell focus-inducing (MCF) viruses derived from AKR-murine leukemia virus (MuLV) are env gene recombinants between ecotropic virus and xenotropic virus. In an attempt to isolate additional env gene recombinants between Friend murine leukemia virus (F-MuLV) and xenotropic virus, we have inoculated cloned ecotropic F-MuLV into newborn NIH Swiss mice and analyzed MuLV released from preleukemic and leukemic spleens of infected mice. Two helper-independent MCF strains of F-MuLV have been isolated. Like the previously described AKR-MCF viruses, the Friend MCF viruses are env gene recombinants between an ecotropic virus (F-MuLV) and a mouse xenotropic virus, as shown by host range, interference pattern, and tryptic peptide analysis of the gp70s of these MuLV. Furthermore, RNA from the Friend MCF viruses hybridizes completely to cDNAsffv, a nucleic acid probe which detects that portion of SFFV which was not derived from P-MuLV. The ability to isolate replicating MCF viruses derived from F-MuLV FURTHER strengthens the parallels between the Friend erythroleukemia system and the AKR thymic leukemia system. Finally, the potential relationship of helper-independent env gene recombinants between F-MuLV and xenotropic virus to be highly leukemogenic SFFV is discussed.

Mapping host range-specific oligonucleotides within genomes of the ecotropic and mink cell focus-inducing strains of Moloney murine leukemia virus.

The site of recombination of a mink cell focus-inducing strain (Mo-MuLV83) derived from an ecotropic Moloney murine leukemia virus (Mo-MuLV) was mapped by fingerprint analysis of the large RNase T1-resistant oligonucleotides, employing a two-dimensional gel electrophoresis method. Mo-MuLV83, in contrast to the ecotropic Mo-MuLV, demonstrated a broadened host range, i.e., growth not only on mouse cells but also on mink cells, and recombination involved the env gene function. The genomic RNA of these two viruses shared 42 out of a total of 51 to 53 large T1 oligonucleotides (81%) and possessed a similar subunit size of 36S. Most of these T1 oligonucleotides were mapped in their relative order to the 3' polyadenylic acid end of the viral RNA molecules. There were 10 common oligonucleotides immediately next to the 3' termini. A cluster of 7 (in Mo-MuLV83) or 10 (in Mo-MuLV) unique T1 oligonucleotides were mapped next to the common sequences at the 3' end, and they all appeared concomitantly in a polyadenylic acid-containing RNA fraction with a sedimentation coefficient slightly larger than 18S. Therefore, the env gene of Mo-MuLV was situated at a location approximately 2,000 to 4,000 nucleotides from the 3' end of the genomic RNA, and the gene order of Mo-MuLV appeared to be similar to that of the more rigorously determined avian oncornaviruses. cDNA(SFFV) specific for the xenotropic sequences in the spleen focus-forming virus RNA hybridized to the cluster of unique oligonucleotides of Mo-MuLV83 RNA. This suggests that the loci of recombination involve the homologous env gene region of a xenotropic virus.

Identification of a 30S RNA with properties of a defective type C virus in murine cells.

A novel species of 30S RNA has been detected in a variety of mouse cell lines. The 30S RNA is specifically packaged by helper-independent type C viruses propagated in such cells. Nucleic acid hybridization detects no homology between the 30SRNA and the genomic RNA of helper-independent mouse type C viruses. The properties of the 30S RNA suggest that it is a defective endogenous mouse type C virus and that it is analogous to a previously described class of defective endogenous rat type C virus, which has been shown previously to be the progenitor of Kirsten and Harvey murine sarcoma viruses.

Friend strain of spleen focus-forming virus is a recombinant between ecotropic murine type C virus and the env gene region of xenotropic type C virus.

The spleen focus-forming virus (SFFV), a replication-defective murine leukemia virus that causes the rapid transformation of certain hematopoietic target cells, has acquired specific xenotropic viral genetic information not contained in Friend helper virus. In the current studies, it is shown that a cDNA that represents a xenotropic virus portion of SFFV detects genetic sequences derived from the env gene region of murine xenotropic virus. The significance of the acquisition of these xenotropic viral sequences by SFFV is discussed with regard to their possible role in the rapid leukemogenicity of SFFV, and an analogy is drawn between the formation of SFFV and the formation of the Kirsten and Harvey sarcoma viruses.

Friend strain of spleen focus-forming virus: a recombinant between mouse type C ecotropic viral sequences and sequences related to xenotropic virus.

The genome of the Friend strain of the spleen focus-forming virus (SFFV) has been analyzed by molecular hybridization. SFFV is composed of genetic sequences homologous to Friend type C helper virus (F-MuLV) and SFFV-specific sequences not present in F-MuLV. These SFFV-specific sequences are present in both the Friend and Rauscher strains of murine erythroleukemia virus. The SFFV-specific sequences are partially homologous to three separate strains of mouse xenotropic virus but not to several cloned mouse ecotropic viruses. Thus, the Friend strain of SFFV appears to be a recombinant between a portion of the F-MuLV genome and RNA sequences that are highly related to murine xenotropic viruses. The implications of the acquisition of the xenotropic virus-related sequences are discussed in relation to the leukemogenicity of SFFV, and a model for the pathogenicity of other murine leukemia-inducing viruses is proposed.