The Fkh1 Forkhead Associated Domain promotes ORC binding to a cohort of DNA replication origins in budding yeast

Binding of chromosomal DNA by the origin recognition complex (ORC) is the pioneering step in DNA replication in eukaryotes. In the budding yeast Saccharomyces cerevisiae, this step is mediated by a specific DNA sequence element within origins, the chromosomal positions that serve as sites of DNA replication initiation. In contrast, metazoan cells rely on multiple features of chromatin to promote ORC-DNA interactions. However, multiple studies reveal tremendous similarities between yeast and metazoan and ORC and provide additional evidence that chromatin also regulates ORC-DNA interactions in this organism. Here, we exploited a distinct cohort of yeast origins that we identified in a previous study where we posited that local origin-adjacent chromatin features promoted normal levels of ORC binding. Specifically, a combination of genetic, genomic and bioinformatic approaches uncovered a role for the Fkh1 protein, and in particular its FHA domain, a phosphothreonine specific binding module, in promoting ORC-origin binding at these yeast origins. The data provide evidence that the Fkh1-FHA domain, acting through a specific Fkh1 binding site in a T-rich orientation located 5' to these origins' ORC binding sites (5' FKH-T site) is required for origin activity. Direct assessment of ORC-origin binding by X-ChIPSeq revealed a strong association between the presence of a 5' FKH-T site and normal levels of ORC binding at this yeast origin cohort and provided evidence for broader association between such an FKH site, the Fkh1-FHA domain and ORC-origin binding across the yeast genome. We propose that the Fkh1-FHA domain is as a non-histone chromatin factors that promotes the normal distribution of ORC-origin DNA complexes over yeast chromosomes