2ND-SITE PROVIRAL ENHANCER ALTERATIONS IN LYMPHOMAS INDUCED BY ENHANCER MUTANTS OF SL3-3 MURINE LEUKEMIA-VIRUS - NEGATIVE EFFECT OF NUCLEARFACTOR-1 BINDING-SITE

SL3-3 is a highly T-lymphomagenic murine retrovirus. Previously, mutat ion of binding sites in the U3 repeat region for the AML1 transcriptio n Factor family (also known as core binding factor [CBF], polyomavirus enhancer binding protein 2 [PEBP2], and SL3-3 enhancer factor 1 [SEF1 ]) were found to strongly reduce the pathogenicity of SW-3 (B. Hallber g, J. Schmidt, A. Luz, F. S. Pedersen, and T. Grundstrom, J. Virol. 65 :4177-4181, 1991). We have now examined the fem cases in which tumors developed harboring proviruses that besides the AML1 (core) site mutat ions carried second-site alterations in their U3 repeat structures. In three distinct cases we observed the same type of alteration which in volved deletions of regions known to contain binding sites for nuclear factor 1 (NF1) and the addition of extra enhancer repeat elements. In transient-expression experiments in T-lymphoid cells, these new U3 re gions acted as stronger enhancers than the U3 regions of the original viruses, This suggests that the altered proviruses represent more-path ogenic variants selected for in the process of tumor formation. To ana lyze the proviral alterations, we generated a series of different enha ncer-promoter reporter constructs, These constructs showed that the ad ditional repeat elements are not critical for enhancer strength, where as the NF1 sites down-regulate the level of transcription in T-lymphoi d cells whether or not the AML1 (core) sites are functional, We theref ore also tested SW-3 viruses with mutated NF1 sites. These viruses hav e unimpaired pathogenic properties and thereby distinguish SW-3 from M oloney murine leukemia virus.