Recombinant retroviruses as a tool for identifying novel oncogenes associated with lymphoma
Description
The recombinant retrovirus, MoFe2, was constructed by replacing the U3 region of Moloney murine leukemia virus (MoMuLV) with homologous sequences from the feline leukemia virus (FeLV-945) LTR. Like other gammaretroviruses, MoMuLV and FeLV induce lymphoma in the natural host through insertional activation of host oncogenes near proviral integrations sites. NIH/Swiss mice neonatally inoculated with MoFe2 developed T-cell lymphomas of immature thymocyte surface phenotype. Southern blot analysis demonstrated that the MoFe2 recombinant did not integrate near the host oncogenes commonly involved in disease induction by either parent virus; thus, MoFe2 represents an opportunity to identify novel oncogenes in the induction of T-cell lymphoma. Common insertion sites (CISs) in MoFe2-induced tumors were identified at six loci, none of which had been previously reported as CISs in tumors induced by either parental virus in wild type animals. Two of the newly identified CISs, 3-19 and Rw1, had not been previously implicated in lymphoma in any retroviral model. The exact function of Rw1 remains unknown and little information is available about the regulation of its expression. The precise location and transcriptional direction of proviral insertions in Rw1 were determined, but proviral insertion demonstrates no clear impact on the level of Rw1 expression in end-stage lymphomas. The results further indicate that Rw1 expression is highly regulated, restricted to a narrow spectrum of cell types, and may be responsive to retroviral infection. The other novel CIS, designated 3-19, encodes the regulatory subunit of phosphatidylinositol 3-kinase gamma (P13Kgamma) termed p101. P13Kgamma is known to be important in a number of cellular functions including chemotaxis, oxidative burst and regulation of T-cell proliferation. P13Kgamma activity is regulated through interactions with G protein-coupled receptors. The fact that p101 is a CIS suggests that its dysregulated expression is associated with T-cell malignancy. Tumors that contained insertions in 3-19 had elevated expression of one or more of the detected transcripts. Overexpression studies revealed that p101 has a positive effect on PI3K by activating Akt, likely by enhancing the activity of p110gamma and sensitizing it to activation. Although overexpression of p101 activates the Akt pathway and does protect cells from DNA damage-induced apoptosis it was also found to induce apoptosis. These findings indicate that substitution of FeLV-945 U3 sequences into the M-MuLV LTR significantly altered the pattern of CIS utilization in the induction of T-cell lymphoma and supports a growing body of evidence that the distinctive sequence and/or structure of the retroviral LTR determines its pattern of insertional activation. These findings demonstrate the utility of the recombinant retrovirus MoFe2-MuLV to in the identification of novel genes implicated in the induction of lymphoma