Cell-Type-Dependent Activity of the Ubiquitous Transcription Factor USF in Cellular Proliferation and Transcriptional Activation

Abstract

USF1 and USF2 are basic helix-loop-helix transcription factors implicated in the control of cellular proliferation. In HeLa cells, the USF proteins are transcriptionally active and their overexpression causes marked growth inhibition. In contrast, USF overexpression had essentially no effect on the proliferation of the Saos-2 osteosarcoma cell line. USF1 and USF2 also lacked transcriptional activity in Saos-2 cells when assayed by transient cotransfection with USF-dependent reporter genes. Yet, there was no difference in the expression, subcellular localization, or DNA-binding activity of the USF proteins in HeLa and Saos-2 cells. Furthermore, Gal4-USF1 and Gal4-USF2 fusion proteins activated transcription similarly in both cell lines. Mutational analysis and domain swapping experiments revealed that the small, highly conserved USF-specific region (USR) was responsible for the inactivity of USF in Saos-2 cells. In HeLa, the USR serves a dual function. It acts as an autonomous transcriptional activation domain at promoters containing an initiator element and also induces a conformational change that is required for USF activity at promoters lacking an initiator. Taken together, these results suggest a model in which the transcriptional activity of the USF proteins, and consequently their antiproliferative activity, is tightly controlled by interaction with a specialized coactivator that recognizes the conserved USR domain and, in contrast to USF, is not ubiquitous. The activity of USF is therefore context dependent, and evidence for USF DNA-binding activity in particular cells is insufficient to indicate USF function in transcriptional activation and growth control.

Loss of USF transcriptional activity in breast cancer cell lines

“…In mutational analysis, the inability of USF to activate transcription in Saos-2 cells correlated with a complete inactivity of the USR domain. These results suggest the existence of a USF-speci®c coactivator that interacts with the USR domain to mediate the transcriptional activity of USF1 and USF2 in cells such as HeLa and is either absent or somehow inactivated in Saos-2 cells (Qyang et al, 1999).…”

“…These cells were used in our experiments at passages 35 to 41. For comparison, two other human cell lines, HeLa (cervical carcinoma) and Saos-2 (osteosarcoma), were added to the panel because they had been previously well characterized for USF expression and activity (Qyang et al, 1999).…”

“…USF2 also contains a second, more classical transcriptional activation domain encoded by the ®fth exon of the gene. Together with the USR, this exon 5 domain is required for transcriptional activation by USF2 of promoters lacking an initiator element (Luo and Sawadogo, 1996b;Qyang et al, 1999). Other USF2 domains, encoded by the ®rst four exons, regulate the activity of the exon 5 activation domain (Luo and Sawadogo, 1996b).…”

“…We recently reported a comparison between the activities of USF1 and USF2 in the HeLa and Saos-2 cell lines (Qyang et al, 1999). In HeLa cells, the USF proteins were transcriptionally active and, when overexpressed, caused marked growth inhibition.…”

The transcription-repression domain of the adenovirus E1A oncoprotein targets p300 at the promoter

“…Gal4-activator plasmids were obtained from several sources. Gal4-NF-kB-p65; Gal4-MyoD (amino acid 1-318); Gal4-cMyb; Gal4-TFE3 (amino acid 2-216) and LexA-RD-ZEB are described in Weintraub et al, 1995 andDean, 1999b; Gal4-USF1 and Gal4-USF2 in Qyang et al, 1999. All plasmids used in transfection studies were purified by double CsCl density gradient centrifugation to facilitate reproducible transfection.…”