THE CARBOXY-TERMINAL TRANSCRIPTION ENHANCEMENT REGION OF THE HUMAN SPUMARETROVIRUS TRANSACTIVATOR CONTAINS DISCRETE DETERMINANTS OF THE ACTIVATOR FUNCTION

The bell gene of human spumaretrovirus (HSRV) encodes a 300-amino-acid nuclear protein termed Bel1 that is a potent activator of transcripti on from the cognate long terminal repeat (LTR). Bell can also efficien tly activate the human immunodeficiency virus type 1 (HIV-1) LTR. We h ave previously shown that the amino-terminal 227-residue region (minim al activator region) of Bell can activate the HSRV LTR at low levels a nd that two distinct domains within the carboxy-terminal 73 residues, from residues 255 to 266 and 272 to 300, that bear little sequence hom ology can independently enhance the activity of the minimal activator domain (L. K. Venkatesh, C. Yang, P. A. Theodorakis, and G. Chinnadura i, J. Virol. 67:161-169, 1993). We now report on the further character ization of these two transcriptional enhancement regions. Mutational a nalysis of the region comprising residues 255 to 266 indicates that a cluster of leucine residues is critical to the function of this region . Also, residues 273 to 287, which are identical in sequence to a 15-a mino-acid segment near the carboxy terminus of the simian foamy virus transcriptional activator Taf, can independently enhance the activity of the minimal activator region. To delineate the region(s) of Bel1 th at could function autonomously as an activator domain, we tested the a ctivity of chimeric proteins comprising either wild-type or functional ly defective forms of Bell fused to the DNA binding domain, Gal4(1-147 ), of the yeast transcriptional activator Gal4 on a synthetic promoter comprising Gal4 DNA binding sites linked to the adenovirus E1B TATA b ox (minimal promoter). Gal4-Bell was found to activate basal transcrip tion from the E1B TATA box at least 35-fold, and the region responsibl e for this activation function was localized to the carboxy-terminal 7 3 amino acids. When the transcriptional enhancement regions were teste d for autonomous activator function as Gal4(1-147) chimeras, residues 272 to 300, but not 255 to 266, were found to activate transcription e fficiently when targeted to the E1B TATA motif and also to HSRV and HI V-1 LTRs. The highly conserved region between amino acids 273 and 287 alone was found to activate transcription efficiently when targeted to the HSRV LTR but not to the E1B TATA box or the HIV-1 LTR. Thus, our results demonstrate that the carboxy-terminal 29-amino-acid region (re sidues 272 to 300) contributes to Bell transactivation by functioning as an autonomous activator of TATA motif-directed transcription in a m anner similar to that of other modular transcriptional activators. In contrast, the domain comprising residues 255 to 266 functions to augme nt transcription only in conjunction with the minimal activator region , which suggests that the two distinct carboxy-terminal activation-enh ancing domains of HSRV Bell may function by different mechanisms.