Regulation of cell cycle transcription factor Swi4 through auto-inhibitionof DNA binding

In Saccharomyces cerevisiae, two transcription factors, SBF (SCB binding fa ctor) and MBF (MCB binding factor), promote the induction of gene expressio n at the G(1)/S-phase transition of the mitotic cell cycle. Swi4 and Mbp1 a re the DNA binding components of SBF and MBF, respectively. The Swi6 protei n is a common subunit of both transcription factors and is presumed to play a regulatory role. SBF binding to its target sequences, the SCBs, is a hig hly regulated event and requires the association of Swi4 with Swi6 through their C-terminal domains. Swi4 binding to SCBs is restricted to the late M and G(1) phases, when Swi6 is localized to the nucleus. We show that in con trast to Swi6, Swi4 remains nuclear throughout the cell cycle. This finding suggests that the DNA binding domain of Swi4 is inaccessible in the full-l ength protein when not complexed with Swi6. To explore this hypothesis, we expressed Swi4 and Swi6 in insect cells by using the baculovirus system. We determined that partially purified Swi4 cannot bind SCBs in the absence of Swi6. However, Swi4 derivatives carrying point mutations or alterations in the extreme C terminus were able to bind DNA or activate transcription in the absence of Swi6, and the C terminus of Swi4 inhibited Swi4 derivatives from binding DNA in trans. Full-length Swi4 was determined to be monomeric in solution, suggesting an intramolecular mechanism for auto-inhibition of binding to DNA by Swi4. We detected a direct in vitro interaction between a C-terminal fragment of Swi4 and the N-terminal 197 amino acids of Swi4, wh ich contain the DNA binding domain. Together, our data suggest that intramo lecular interactions involving the C-terminal region of Swi4 physically pre vent the DNA binding domain from binding SCBs. The interaction of the carbo xy-terminal region of Swi4 with Swi6 alleviates this inhibition, allowing S wi4 to bind DNA.