A hallmark of histone H3 lysine 9 (H3K9) methylated heterochromatin, conserved from fission yeast,Schizosaccharomyces pombe (S. pombe), to humans, is its ability to spread to adjacent genomic regions1–6. Central to heterochromatin spread is the heterochromatin protein 1 (HP1), which recognizes H3K9 methylated chromatin, oligomerizes, and forms a versatile platform that participates in diverse nuclear functions, ranging from gene silencing to chromosome segregation1–6. How HP1 proteins assemble on methylated nucleosomal templates and how the HP1-nucleosome complex achieves functional versatility remain poorly understood. Here, we show that binding of the major S. pombe HP1 protein, Swi6, to methylated nucleosomes drives a switch from an auto-inhibited state to a spreading competent state. In the auto-inhibited state, a histone mimic sequence in one Swi6 monomer blocks methyl mark recognition by the chromodomain of another monomer. Auto-inhibition is relieved by recognition of two template features, the H3K9 methyl mark and nucleosomal DNA. Cryo-Electron Microscopy (EM) based reconstruction of the Swi6-nucleosome complex provides the overall architecture of the spreading-competent state in which two unbound chromodomain sticky ends appear exposed. Disruption of the switch between the auto-inhibited and spreading competent state disrupts heterochromatin assembly and gene silencing in vivo. These findings are reminiscent of other conditionally activated polymerization processes, such as actin nucleation, and open up a new class of regulatory mechanisms that operate on chromatin in vivo.

Long noncoding RNAs (lncRNAs) can trigger repressive chromatin, but how they recruit silencing factors remains unclear. In Schizosaccharomyces pombe, heterochromatin assembly on transcribed noncoding pericentromeric repeats requires both RNAi and RNAi-independent mechanisms. In Saccharomyces cerevisiae, which lacks a repressive chromatin mark (H3K9me [methylated Lys9 on histone H3]), unstable ncRNAs are recognized by the RNA-binding protein Nrd1. We show that the S. pombe ortholog Seb1 is associated with pericentromeric lncRNAs. Individual mutation of dcr1 + (Dicer) or seb1 + results in equivalent partial reductions of pericentromeric H3K9me levels, but a double mutation eliminates this mark. Seb1 functions independently of RNAi by recruiting the NuRD (nucleosome remodeling and deacetylase)-related chromatin-modifying complex SHREC (Snf2-HDAC [histone deacetylase] repressor complex).Supplemental material is available for this article.Received November 1, 2012; revised version accepted August 7, 2013. A major unsolved question in chromatin biology is how long intergenic noncoding RNAs (lincRNAs) trigger the formation of repressed chromatin. A large number of mammalian lincRNAs have been identified by systematic studies ). Many of these ncRNAs associate with chromatin-modifying complexes (Khalil et al. 2009). Multiple models have been proposed for how these ncRNAs are recognized and recruit chromatinmodifying factors, but little is understood mechanistically (Guttman and Rinn 2012).In Schizosaccharomyces pombe, pericentromeric heterochromatin assembly is promoted by transcription of the dg and dh repeat sequences by RNA polymerase II (Pol II) (Djupedal et al. 2005;Kato et al. 2005). The corresponding long ncRNAs (lncRNAs) are converted into dsRNAs and processed into siRNAs by the combined action of RNA-directed RNA polymerase complex (RDRC) and Dicer (Dcr1) (Verdel et al. 2009;Lejeune and Allshire 2011). siRNAs produced by Dicer are bound by Argonaute (Ago1), a component of the RNA-induced transcriptional silencing (RITS) complex, and together they promote both degradation of pericentromeric ncRNAs and transcriptional silencing via repressive histone methylation (Verdel et al. 2004). These complexes in turn recruit the Clr4 methyltransferase complex (CLRC), which methylates Lys9 on histone H3 (H3K9me) (Nakayama et al. 2001;Zhang et al. 2008). The methyl mark serves as a binding platform for the repressive HP1 proteins Swi6 and Chp2 (Thon and Verhein-Hansen 2000;Bannister et al. 2001;Fischer et al. 2009). Both proteins promote the recruitment of SHREC (Snf2-HDAC [histone deacetylase] repressor complex) to pericentromeric heterochromatin (Sugiyama et al. 2007;Sadaie et al. 2008). Moreover, Chp2 has been found to associate with SHREC to form the SHREC2 complex (SHREC complex associated with Chp2) (Motamedi et al. 2008). The core of SHREC consists of silencing factors Clr1 and Clr2, the HDAC Clr3, and the putative chromatin-remodeling enzyme Mit1 (Sugiyama et al. 2007). SHREC and SHREC2 resemble the mammalian nucle...

The biological risks of the deep space environment must be elucidated to enable a new era of human exploration and scientific discovery beyond low earth orbit (LEO). There is a paucity of deep space biological missions that will inform us of the deleterious biological effects of prolonged exposure to the deep space environment. To safely undertake long-term missions to Mars and space habitation beyond LEO, we must first prove and optimize autonomous biosensors to query the deep space radiation environment. Such biosensors must contain organisms that can survive for extended periods with minimal life support technology and must function reliably with intermittent communication with Earth. NASA's BioSentinel mission, a nanosatellite containing the budding yeast Saccharomyces cerevisiae, is such a biosensor and one of the first biological missions beyond LEO in nearly half a century. It will help fill critical gaps in knowledge about the effects of uniquely composed, chronic, low-flux deep space radiation on biological systems and in particular will provide valuable insight into the DNA damage response to highly ionizing particles. Due to yeast's robustness and desiccation tolerance, it can survive for periods analogous to that of a human Mars mission. In this study, we discuss our optimization of conditions for long-term reagent storage and yeast survival under desiccation in preparation for the BioSentinel mission. We show that long-term yeast cell viability is maximized when cells are air-dried in trehalose solution and stored in a low-relative humidity and low-temperature environment and that dried yeast is sensitive to low doses of deep space-relevant ionizing radiation under these conditions. Our findings will inform the design and development of improved future long-term biological missions into deep space.

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