Fkh1 and Fkh2 Bind Multiple Chromosomal Elements in the S. cerevisiae Genome with Distinct Specificities and Cell Cycle Dynamics

A Zachary Ostrow,Catherine A Fox,Simon Tavaré,Tittu Nellimoottil,Simon R V Knott,Oscar M Aparicio,Yanchang Wang

doi:10.1371/journal.pone.0087647

A Zachary Ostrow, Catherine A Fox + Show 5 more

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https://doi.org/10.1371/journal.pone.0087647

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Abstract

Forkhead box (FOX) transcription factors regulate a wide variety of cellular functions in higher eukaryotes, including cell cycle control and developmental regulation. In Saccharomyces cerevisiae, Forkhead proteins Fkh1 and Fkh2 perform analogous functions, regulating genes involved in cell cycle control, while also regulating mating-type silencing and switching involved in gamete development. Recently, we revealed a novel role for Fkh1 and Fkh2 in the regulation of replication origin initiation timing, which, like donor preference in mating-type switching, appears to involve long-range chromosomal interactions, suggesting roles for Fkh1 and Fkh2 in chromatin architecture and organization. To elucidate how Fkh1 and Fkh2 regulate their target DNA elements and potentially regulate the spatial organization of the genome, we undertook a genome-wide analysis of Fkh1 and Fkh2 chromatin binding by ChIP-chip using tiling DNA microarrays. Our results confirm and extend previous findings showing that Fkh1 and Fkh2 control the expression of cell cycle-regulated genes. In addition, the data reveal hundreds of novel loci that bind Fkh1 only and exhibit a distinct chromatin structure from loci that bind both Fkh1 and Fkh2. The findings also show that Fkh1 plays the predominant role in the regulation of a subset of replication origins that initiate replication early, and that Fkh1/2 binding to these loci is cell cycle-regulated. Finally, we demonstrate that Fkh1 and Fkh2 bind proximally to a variety of genetic elements, including centromeres and Pol III-transcribed snoRNAs and tRNAs, greatly expanding their potential repertoire of functional targets, consistent with their recently suggested role in mediating the spatial organization of the genome.

Highlights

Forkhead Box (Fox) transcription factors comprise a large and diversified family of DNA binding proteins that function in a wide range of processes from yeast to humans, including cell cycle control, development, stress response, and apoptosis
To examine the Fkh1- and Fkh2-bound loci we have identified in relation to replication origins, we divided origins into three groups defined by their change in origin activity in fkh1D fkh2D cells in our previous study: Fkhactivated origins, which showed reduced early firing, Fkhrepressed origins, which showed increased early firing, and Fkhunregulated origins, which showed no significant change in early firing [24]
The results show no instances of Mcm1 binding sites within 500 bp of any of the replication origin classes, whereas 19% of CLB2-cluster genes are within 500 bp of an Mcm1 binding site (Fig. 5C)

Summary

Introduction

Forkhead Box (Fox) transcription factors comprise a large and diversified family of DNA binding proteins that function in a wide range of processes from yeast to humans, including cell cycle control, development, stress response, and apoptosis (reviewed in [1]). Common to these proteins is the Forkhead DNA-Binding Domain (Fkh-DBD) that binds DNA as a monomer through a conserved helix-turn-helix motif variant, known as a winged-helix. Fkh and Fkh regulate a set of ,33 genes, referred to as the CLB2cluster, which are expressed during late S/G2-phase to regulate subsequent mitotic events [11]

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