Alternative arrangements of telomeric recognition sites regulate the binding mode of the DNA-binding domain of yeast Rap1

The function of yeast Rap1 as an activator in transcription, a repressor at silencer elements, and as a major component of the shelterin-like complex at telomeres requires the known high-affinity and specific interaction of the DNA-binding domain (DBD) with its recognition sequences. In addition to a high-affinity one-to-one complex with its DNA recognition site, Rap1^DBD also forms lower affinity complexes with higher stoichiometries on DNA. We proposed that this originates from the ability of Rap1^DBD to access at least two DNA-binding modes. In this work, we show that Rap1^DBD binds in multiple binding modes to recognition sequences that contain different spacer lengths between the hemi-sites. We also provide evidence that in the singly-ligated complex Rap1^DBD binds quite differently to these sequences. Rap1^DBD also binds to a single half-site but does so using the alternative DNA-binding mode where only a single Myb-like domain interacts with DNA. We found that all arrangements of Rap1 sites tested are represented within the telomeric sequence and our data suggest that at telomeres Rap1 might form a nucleoprotein complex with a heterogeneous distribution of bound states.