GEOapplication/xmlftp://ftp.ncbi.nlm.nih.gov/geo/series/GSE346nnn/GSE346150/primaryOK200GenomicsSaccharomyces cerevisiaeGenome binding/occupancy profiling by high throughput sequencinghttps://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE346150GEOGSEfalseATAC-seq and MNase-seq Detect Distinct Modes of Chromatin Accessibility [Mnase]Chromatin accessibility shapes the ability of transcription factors (TFs) and the transcriptional machinery to engage genomic DNA and therefore plays a central role in gene regulation. Two widely used approaches for profiling chromatin accessibility are micrococcal nuclease (MNase)-seq and assay for transposase-accessible chromatin (ATAC)-seq. ATAC-seq peaks are often thought to be equivalent to nucleosome-depleted regions (NDRs) that are defined by MNase-seq; however, these two measurements have not been systematically compared. Here, we performed a side-by-side comparison of ATAC-seq and MNase-seq in budding yeast and found a substantial discrepancy between ATAC-seq peaks and MNase-defined NDRs. This discrepancy is not primarily explained by intrinsic differences between MNase and Tn5 enzymatic activity. Instead, ATAC-seq peaks and NDRs capture distinct chromatin states. Specifically, ATAC-seq peaks are enriched at dynamic nucleosomes associated with transcriptional co-regulators, including SAGA and SWI/SNF, whereas NDRs mark more static nucleosome-free regions at promoters. Depletion of SWI/SNF, but not RSC, reduces ATAC-seq signals. Generation of NDRs and ATAC-seq peaks requires distinct TF properties, and native TFs differ in their ability to produce these two types of open chromatin. Finally, we show that the functional distinction between ATAC-seq peaks and NDRs are widespread across eukaryotic species, including human cells. Together, our results provide new insights into the biological meaning of chromatin accessibility measured by these two assays.2026/09/08GSE346150GSM10025297GSM10025296GSM10025299GSM10025298GSM10025301GSM1002530031112346150Saccharomyces cerevisiae