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| Status |
Public on Sep 11, 2024 |
| Title |
Heterogeneous single-molecule packaging of human mitochondrial nucleoids |
| Organism |
Drosophila melanogaster |
| Experiment type |
Other
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| Summary |
During eukaryotic transcription, RNA polymerases must initiate and pause within a crowded, complex environment, surrounded by nucleosomes and other transcriptional activity. This environment creates a spatial arrangement along individual chromatin fibers ripe for both competition and coordination, yet these relationships remain largely unknown owing to the inherent limitations of traditional structural and sequencing methodologies. To address these limitations, we employed long-read chromatin fiber sequencing (Fiber-seq) to visualize RNA polymerases within their native chromatin context at single-molecule and near single-nucleotide resolution along up to 30 kb fibers. We demonstrate that Fiber-seq enables the identification of single-molecule RNA Polymerase (Pol) II and III transcription associated footprints, which, in aggregate, mirror bulk short-read sequencing-based measurements of transcription. We show that Pol II pausing destabilizes downstream nucleosomes, with frequently paused genes maintaining a short-term memory of these destabilized nucleosomes. Furthermore, we demonstrate pervasive direct coordination and anti-coordination between nearby Pol II genes, Pol III genes, transcribed enhancers, and insulator elements. This coordination is largely limited to spatially organized elements within 5 kb of each other, implicating short-range chromatin environments as a predominant determinant of coordinated polymerase initiation. Overall, transcription initiation reshapes surrounding nucleosome architecture and coordinates nearby transcriptional machinery along individual chromatin fibers.
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| Overall design |
PacBio sequencing of exogenously methylated genomic DNA from Drosophila S2 cells
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| Contributor(s) |
Tullius TW, Isaac SR, Ranchalis J, Dubocanin D, Stergachis AB, Churchman SL |
| Citation(s) |
39191261 |
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| Submission date |
Dec 08, 2023 |
| Last update date |
Sep 12, 2024 |
| Contact name |
Thomas Tullius |
| E-mail(s) |
tommytullius@mac.com
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| Organization name |
Harvard Medical School
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| Department |
Genetics
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| Lab |
Stirling Churchman
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| Street address |
77 Ave Louis Pasteur
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
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| Platforms (1) |
| GPL28353 |
Sequel II (Drosophila melanogaster) |
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| Samples (12)
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| Relations |
| BioProject |
PRJNA1050275 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE249742_RAW.tar |
6.7 Gb |
(http)(custom) |
TAR (of BED) |
SRA Run Selector |
| Raw data are available in SRA |
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