Long-Range Effects of Retroviral Insertion on c -myb : Overexpression May Be Obscured by Silencing during Tumor Growth In Vitro

Abstract

ABSTRACT The c -myb oncogene is a frequent target for retroviral activation in hemopoietic tumors of avian and mammalian species. While insertions can target the gene directly, numerous clusters of retroviral insertion sites have been identified which map close to c -myb and outside the transcription unit in T-lymphomas ( Ahi-1 , fit-1 , and Mis-2 ) and monocytic and myeloid leukemias ( Mml1 , Mml2 , Mml3 , and Epi-1 ). Previous analyses showed no consistent effect of these insertions on c -myb expression, raising the possibility that other nearby genes were the true targets. In contrast, our analysis of four cell lines established from lymphomas bearing insertions at fit-1 ( fti-1 ) (feline leukemia virus) and Ahi-1 (Moloney murine leukemia virus) shows that these display higher expression levels of c -myb RNA and protein compared to a panel of phenotypically similar cell lines lacking such insertions. An interesting feature of the cell lines with long-range c- myb insertions was that each also carried an activated Myc allele. The potential for oncogenic synergy between Myb and Myc in T-cell lymphoma was confirmed in transgenic mice overexpressing alleles of both genes in the T-cell compartment, lending further credence to the case for c- myb as the major target for long-range activation. In contrast, mapping and analysis of c- myb neighboring genes ( HBS1 and FLJ20069 ) showed that the expression of these genes did not correlate well with the presence of proviral insertions. A possible explanation for the paradoxical behavior of c -myb was provided by one of the murine T-lymphoma lines bearing an insertion at Ahi-1 (p/m16i) that reproducibly down-regulated c -myb RNA and protein to very low levels or undetectable levels on prolonged culture. Our observations implicate c -myb as a key target of upstream and downstream retroviral insertions. However, overexpression may become dispensable during outgrowth in vitro, and perhaps during tumor progression in vivo, providing a potential rationale for the previously observed discordance between retroviral insertion and c -myb expression levels.