SummaryNucleosome organization has a key role in transcriptional regulation, yet the precise mechanisms establishing nucleosome locations and their effect on transcription are unclear. Here, we use an induced degradation system to screen all yeast ATP-dependent chromatin remodelers. We characterize how rapid clearance of the remodeler affects nucleosome locations. Specifically, depletion of Sth1, the catalytic subunit of the RSC (remodel the structure of chromatin) complex, leads to rapid fill-in of nucleosome-free regions at gene promoters. These changes are reversible upon reintroduction of Sth1 and do not depend on DNA replication. RSC-dependent nucleosome positioning is pivotal in maintaining promoters of lowly expressed genes free from nucleosomes. In contrast, we observe that upon acute stress, the RSC is not necessary for the transcriptional response. Moreover, RSC-dependent nucleosome positions are tightly related to usage of specific transcription start sites. Our results suggest organizational principles that determine nucleosome positions with and without RSC and how these interact with the transcriptional process.

Background: Spore germination in the yeast Saccharomyces cerevisiae is a process in which nondividing haploid spores re-enter the mitotic cell cycle and resume vegetative growth. To study the signals and pathways underlying spore germination we examined the global changes in gene expression and followed cell-cycle and germination markers during this process.

Background: Meiosis in budding yeast is coupled to the process of sporulation, where the four haploid nuclei are packaged into a gamete. This differentiation process is characterized by a point of transition, termed commitment, when it becomes independent of the environment. Not much is known about the mechanisms underlying commitment, but it is often assumed that positive feedback loops stabilize the underlying gene-expression cascade.

SummaryTranscription factor (TF) binding to DNA is crucial for transcriptional regulation. There are multiple methods for mapping such binding. These methods balance between input requirements, spatial resolution, and compatibility with high-throughput automation. Here, we describe SLIM-ChIP (short-fragment-enriched, low-input, indexed MNase ChIP), which combines enzymatic fragmentation of chromatin and on-bead indexing to address these desiderata. SLIM-ChIP reproduces a high-resolution binding map of yeast Reb1 comparable with existing methods, yet with less input material and full compatibility with high-throughput procedures. We demonstrate the robustness and flexibility of SLIM-ChIP by probing additional factors in yeast and mouse. Finally, we show that SLIM-ChIP provides information on the chromatin landscape surrounding the bound transcription factor. We identify a class of Reb1 sites where the proximal −1 nucleosome tightly interacts with Reb1 and maintains unidirectional transcription. SLIM-ChIP is an attractive solution for mapping DNA binding proteins and charting the surrounding chromatin occupancy landscape at a single-cell level.

In the last decade, multiple studies demonstrated that cells maintain a balance of mRNA production and degradation, but the mechanisms by which cells implement this balance remain unknown. Here, we monitored cells’ total and recently-transcribed mRNA profiles immediately following an acute depletion of Xrn1—the main 5′-3′ mRNA exonuclease—which was previously implicated in balancing mRNA levels. We captured the detailed dynamics of the adaptation to rapid degradation of Xrn1 and observed a significant accumulation of mRNA, followed by a delayed global reduction in transcription and a gradual return to baseline mRNA levels. We found that this transcriptional response is not unique to Xrn1 depletion; rather, it is induced earlier when upstream factors in the 5′-3′ degradation pathway are perturbed. Our data suggest that the mRNA feedback mechanism monitors the accumulation of inputs to the 5′-3′ exonucleolytic pathway rather than its outputs.