Spi-1/PU.1 is a positive regulator of the Fli-1 gene involved in inhibition of erythroid differentiation in Friend erythroleukemic cell lines

Spi-1/PU.1 and Fli-1 are two members of the ETS family of transcription fac tors whose expression is deregulated by proviral insertion in most erythrol eukemic cell lines induced by the spleen focus-forming virus (SFFV) and Fri end murine leukemia virus (F-MuLV) components of the Friend viral complex, respectively. In this study, we present evidence that transcription of the Fli-1 gene is positively regulated by Spi-1/PU.1 in SFFV-transformed cell l ines: (i) all SFFV-transformed cell lines expressing Spi-1/PU.1 are charact erized by a specific pattern of Fli-1 gene transcripts initiated in the -20 0 region instead of position -400 as reported for F-MuLV-transformed cell l ines; (ii) these Fli-1 transcripts initiated in the -200 region are downreg ulated in parallel with that of Spi-1/PU.1 during hexamethylenebisacetamide (HMBA) induced differentiation; and (iii) Fli-1 transcription is upregulat ed in SFFV cells lines following stable transfection of a Spi-1/PU.1 expres sion vector. Furthermore, we found by transient transfection assays that th e -270/-41 region of the Fli-1 gene displays promoter activity which is tra nsactivated by Spi-1/PU.1. This promoter is strictly dependent on the integ rity of two highly conserved ETS DNA binding sites that bind the Spi-1/PU.1 protein in vitro. Finally, we show that transfection of constitutive or in ducible Fli-1 expression vectors in SFFV-transformed cells inhibits their e rythroid differentiation induced by HMBA. Overall, these data indicate that Fli-1 is a target gene of the Spi-1/PU.1 transcription factor in SFFV-tran sformed cell lines. We further suggest that deregulated synthesis of Fli-1 may trigger a common mechanism contributing to erythroleukemia induced by e ither SFFV or F-MuLV.