Murine leukemia virus proviral insertions between the N-ras and unr genes in B-cell lymphoma DNA affect the expression of N-ras only.
Akv1-99, a variant of Akv murine leukemia virus, induces B-cell lymphomas with nearly 100% incidence and a mean latency period of 12 months after injection into newborn NMRI mice. PCR amplification and sequence analyses of DNA flanking integrated proviruses revealed proviral insertion into the N-ras/unr (upstream of N-ras) locus in 2 out of 13 B-cell lymphomas, both of which appeared clonal by Southern blotting analysis. These two tumors showed increased expression levels of N-ras by Northern blotting, as did a third tumor shown by reverse transcriptase PCR to have a nonclonal provirus integration located in the same area. However, no significant changes in expression were observed when using a specific probe for the unr gene. All proviruses were integrated in the same transcriptional orientation as unr and N-ras genes. By promoter insertion, the two Akv1-99 proviruses integrated between exon -1 and exon 1 of N-ras gave rise to two different spliced products, whereas the provirus integrated into unr used only an exon skipping pattern. The absence of mutations of the N-ras codons 12, 13, 18, and 61 suggests that activation of the proto-oncogene is exclusively due to overexpression by retroviral promoter insertion, and furthermore, Northern blot analyses indicate that the expression of unr is unaffected by N-ras overexpression even in the case where the unr gene itself is the target of proviral insertion. Thus, altogether our findings indicate that overexpression of N-ras plays a role in development of murine leukemia virus-induced B-cell lymphomas, leaving the expression of the tightly linked unr gene unaltered.
Martín-Hernández
J
J
Department of Molecular and Structural Biology, University of Aarhus, DK-8000 Aarhus C, Denmark.
Sørensen
A B
AB
Pedersen
F S
FS
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D017931:DNA Primers; D004273:DNA, Neoplasm / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000502:physiology; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D011905:Genes, ras; D009052:Leukemia Virus, Murine / Q000235:genetics*; D016393:Lymphoma, B-Cell / Q000235:genetics*; D051379:Mice; D018345:Mice, Knockout; D017354:Point Mutation; D000090063:Proto-Oncogene Mas; D011533:Proviruses / Q000235:genetics*; D012326:RNA Splicing; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction
C079950:CSDE1 protein, human; D017931:DNA Primers; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D016601:RNA-Binding Proteins