Centromere RNA Is Negatively Regulated by Cbf1 and Its Unscheduled Synthesis Impacts CenH3 Binding. Author Chi-Fu Chen, Thomas Pohl, Angela Chan, Joshua Slocum, Virginia Zakian Publication Year 2019 Type Journal Article Abstract Two common features of centromeres are their transcription into noncoding centromere RNAs (cen-RNAs) and their assembly into nucleosomes that contain a centromere-specific histone H3 (cenH3). Here, we show that cen-RNA was present in low amounts in wild-type (WT) cells, and that its appearance was tightly cell cycle-regulated, appearing and disappearing in a narrow window in S phase after centromere replication. In cells lacking Cbf1, a centromere-binding protein, cen-RNA was 5-12 times more abundant throughout the cell cycle. In WT cells, cen-RNA appearance occurred at the same time as loss of Cbf1's centromere binding, arguing that the physical presence of Cbf1 inhibits cen-RNA production. Binding of the Pif1 DNA helicase, which happens in mid-late S phase, occurred at about the same time as Cbf1 loss from the centromere, suggesting that Pif1 may facilitate this loss by its known ability to displace proteins from DNA. Cen-RNAs were more abundant in Δ cells but only in mid-late S phase. However, fork pausing at centromeres was not elevated in Δ cells but rather was due to centromere-binding proteins, including Cbf1 Strains with increased cen-RNA lost centromere plasmids at elevated rates. In Δ cells, where both the levels and the cell cycle-regulated appearance of cen-RNA were disrupted, the timing and levels of cenH3 centromere binding were perturbed. Thus, cen-RNAs are highly regulated, and disruption of this regulation correlates with changes in centromere structure and function. Keywords RNA, Untranslated, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Chromosomal Proteins, Non-Histone, Chromosome Segregation, Kinetochores, RNA, Fungal, Histones, Nucleosomes, Chromatin, DNA Helicases, Centromere, Basic Helix-Loop-Helix Leucine Zipper Transcription Factors Journal Genetics Volume 213 Issue 2 Pages 465-479 Date Published 2019 Oct ISSN Number 1943-2631 DOI 10.1534/genetics.119.302528 Alternate Journal Genetics PMCID PMC6781895 PMID 31391265 PubMedPubMed CentralGoogle ScholarBibTeXEndNote X3 XML