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Synthesis of ribosomal RNAs (rRNAs) is the major transcriptional event in proliferating cells. In eukaryotes, ribosomal DNA (rDNA) is transcribed by RNA polymerase I from a multicopy locus coexisting in at least two different chromatin states. This heterogeneity of rDNA chromatin has been an obstacle to defining its molecular composition. We developed an approach to analyze differential protein association with each of the two rDNA chromatin states in vivo in the yeast Saccharomyces cerevisiae. We demonstrate that actively transcribed rRNA genes are largely devoid of histone molecules, but instead associate with the high-mobility group protein Hmo1.[Keywords: Chromatin; transcription; ribosomal DNA] Supplemental material is available at http://www.genesdev.org. Received December 11, 2007; revised version accepted February 29, 2008. In the yeast Saccharomyces cerevisiae (hereafter called yeast), >60% of total cellular transcripts are produced from the ribosomal DNA (rDNA) locus by a specific multiprotein enzyme, RNA Polymerase I (Pol I), and an assorted set of additional transcription factors (Reeder 1999;Moss and Stefanovsky 2002;Russell and Zomerdijk 2006). The basal Pol I transcription machinery has been largely defined and consists of two multiprotein complexes, the upstream activating factor (UAF) , and the core factor (CF) (Keys et al. 1994), as well as the TATA-box-binding protein (TBP) (Steffan et al. 1996).The template of RNA Pol I transcription, the yeast ribosomal RNA (rRNA) genes are located on the right arm of chromosome XII in a tandem array of 150-200 copies (Fig. 1), representing almost 10% of the yeast genome. Each of the repeats contains the Pol I-transcribed 35S rRNA gene (precusor of the 5.8S, 18S, and 25S rRNAs) and the RNA Polymerase III (Pol III)-transcribed 5S rRNA gene (Fig. 1). Three different regulatory DNA elements have been identified within the 35S rRNA gene sequence, two of which are located at the 5Ј end of the 35S rDNA, spanning only ∼170 base pairs (bp) (Musters et al. 1989;Kulkens et al. 1991): the upstream element (UE), which is the binding site for the UAF; and the core promoter (CP), where CF can be recruited (Fig. 1). The third element, called the enhancer (ENH), is located at the 3Ј end of the 35S transcription unit (Fig. 1) and was originally identified as exhibiting a stimulatory effect on Pol I transcription in experiments using episomal reporter systems Warner 1984, 1986). However, this sequence has been shown subsequently to be dispensable for RNA Pol I transcription in the chromosomal context (Wai et al. 2001).In each cell, Pol I-transcribed rDNA coexists in two different chromatin states (Conconi et al. 1989). In these studies, cells were treated with the DNA cross-linking drug psoralen, and it was shown that actively transcribed and transcriptionally inactive rRNA genes differed substantially in their degree of psoralen incorporation. The different migration behavior of the corresponding restriction fragments in agarose gel electrophoresis led to the terms s-ban...

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Translational repression and mRNA degradation are critical mechanisms of posttranscriptional gene regulation that help cells respond to internal and external cues. In response to certain stress conditions, many mRNA decay factors are enriched in processing bodies (PBs), cellular structures involved in degradation and/or storage of mRNAs. Yet, how cells regulate assembly and disassembly of PBs remains poorly understood. Here, we show that in budding yeast, mutations in the DEAD-box ATPase Dhh1 that prevent ATP hydrolysis, or that affect the interaction between Dhh1 and Not1, the central scaffold of the CCR4-NOT complex and an activator of the Dhh1 ATPase, prevent PB disassembly in vivo. Intriguingly, this process can be recapitulated in vitro, since recombinant Dhh1 and RNA, in the presence of ATP, phase-separate into liquid droplets that rapidly dissolve upon addition of Not1. Our results identify the ATPase activity of Dhh1 as a critical regulator of PB formation.DOI: http://dx.doi.org/10.7554/eLife.18746.001