An Abf1p C-terminal region lacking transcriptional activation potential stimulates a yeast origin of replication

Abstract

Although it has been demonstrated that eukaryotic cellular origins of DNA replication may harbor stimulatory elements that bind transcription factors, how these factors stimulate origin function is unknown. In Saccharomyces cerevisiae, the transcription factor Abf1p stimulates origin function of ARS121 and ARS1. In the results presented here, an analysis of Abf1p function has been carried out utilizing LexA(BD)-Abf1p fusion proteins and an ARS121 derivative harboring LexA DNA-binding sites. A minimal region which stimulates origin function mapped to 50 amino acids within the C-terminus of Abf1p. When tested for transcriptional activation of a LacZ reporter gene, the same LexA(BD)-Abf1p fusion protein had negligible transcriptional activation potential. Therefore, stimulation of ARS121 may occur independently of a transcriptional activation domain. It has been previously observed that the Gal4p, Rap1p DNA-binding sites and the LexA-Gal4p fusion protein can replace the role of Abf1p in stimulating ARS1. Here we show that the stimulatory function of Abf1p at ARS121 cannot be replaced by these alternative DNA-binding sites and the potent chimeric transcriptional activator LexA(BD)-Gal4(AD)p. Hence, these results strongly suggest that the Abf1p stimulation of replication may differ for ARS121 and ARS1, and imply specificity in the Abf1p/ARS121 relationship.