Reb1, Cbf1, and Pho4 Bias Histone Sliding and Deposition Away from Their Binding Sites

ABSTRACT

In transcriptionally active genes, nucleosome positions in promoters are regulated by nucleosome-displacing factors (NDFs) and chromatin-remodeling enzymes. Depletion of NDFs or the RSC chromatin remodeler shrinks or abolishes the nucleosome-depleted regions (NDRs) in promoters, which can suppress gene activation and result in cryptic transcription. Despite their vital cellular functions, how the action of chromatin remodelers may be directly affected by site-specific binding factors like NDFs is poorly understood. Here, we demonstrate that two NDFs, Reb1 and Cbf1, can direct both Chd1 and RSC chromatin-remodeling enzymes in vitro, stimulating repositioning of the histone core away from their binding sites. Interestingly, although the Pho4 transcription factor had a much weaker effect on nucleosome positioning, both NDFs and Pho4 were able to similarly redirect positioning of hexasomes. In chaperone-mediated nucleosome assembly assays, Reb1 but not Pho4 showed an ability to block deposition of the histone H3/H4 tetramer, but Reb1 did not block addition of the H2A/H2B dimer to hexasomes. Our in vitro results show that NDFs bias the action of remodelers to increase the length of the free DNA in the vicinity of their binding sites. These results suggest that NDFs could directly affect NDR architecture through chromatin remodelers.

KEYWORDS:

• centromere-binding factor 1 (Cbf1)
• chromatin remodeling
• chromodomain helicase DNA-binding protein 1 (Chd1)
• nucleosome-depleted region (NDR)
• nucleosome-displacing factor (NDF)
• RNA polymerase I enhancer binding protein 1 (Reb1)
• remodels the structure of chromatin (RSC)
• hexasome
• nucleosome

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Declaration of Interests

The authors declare no conflict of interest.

SUPPLEMENTAL MATERIAL

Supplemental material is available online only.

ACKNOWLEDGMENTS

We thank Michael Poirier for providing Pho4 and Cbf1 proteins, the Reb1 expression vector, and ΔR and R⁷⁻ templates, and Carl Wu for providing RSC complex.

This work was supported by the National Institutes of Health grant R01GM084192.

S.G.K. conceptualized the study, carried out the experiments, wrote the original draft and revised the manuscript. G.D.B. conceptualized the study, revised the manuscript, and received the funding.

We declare no conflict of interest.