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| Status |
Public on Nov 24, 2010 |
| Title |
Silencing factors induce the elimination of nucleosome-free regions in S. pombe heterochromatin |
| Organism |
Schizosaccharomyces pombe |
| Experiment type |
Expression profiling by genome tiling array
Genome binding/occupancy profiling by genome tiling array
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| Summary |
Abstract: In the yeast S. pombe, multiple chromatin-modifying enzymes are required for heterochromatin formation, yet how their actions alter chromatin structure to block access to DNAby the transcriptional machinery is unknown. We constructed high-resolution nucleosome occupancy maps of heterochromatic regions in wild-type strains and in mutants lacking the H3K9 methyltransferase Clr4 or one of the two activities of the silencing effector complex SHREC. Fourier analysis reveals that these enzymes do not increase the regularity of nucleosome spacing. Rather, their principal effect was to induce the elimination of nucleosome-free regions (NFRs). Both NFRs associated with transcription initiation sites as well as those not associated with promoters are affected. As in S. cerevisiae, repressed genes in euchromatin retain their NFRs. Thus, NFR elimination cannot be explained as a secondary consequence of repression. The maps also show that TFIIIC boundary elements have NFRs resistant to silencing, suggesting a potential
role in preventing lateral spread of heterochromatin. NFR elimination offers a mechanism by which heterochromatin restricts access of the transcriptional machinery to DNA.
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| Overall design |
Nucleosome positions in either heterochromatin or euchromatin were mapped in various mutants to determine whether chromatin structure is altered upon heterochromatic silencing. Eight strains were assayed for nucleosome positioning in heterochromatin while two strains were assayed for nucleosome positioning in euchromatin. Two biological replicates were assayed for each strain. No dye swaps were done.
RNA transcripts were also mapped using the heterochromatin tiling arrays to determine where RNA transcripts accumulate in heterochromatin. Three strains were assayed for transcripts in heterochromatin by hybridizing cy5-labeled cDNA created from total RNA against cy3-labeled cDNA created from RNA synthesized from sonicated genomic DNA. Two biological replicates were assayed for each strain.
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| Contributor(s) |
Garcia JF, Dumesic PA, Hartley PD, El_Samad H, Madhani HD |
| Citation(s) |
20675407 |
| Submission date |
Dec 21, 2009 |
| Last update date |
Mar 21, 2012 |
| Contact name |
Hiten Madhani |
| E-mail(s) |
hitenmadhani@gmail.com
|
| URL |
http://madhanilab.ucsf.edu
|
| Organization name |
University of California, San Francisco
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| Department |
Biochemistry and Biophysics
|
| Street address |
600 16th St.
|
| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94143-2200 |
| Country |
USA |
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| Platforms (2) |
| GPL9819 |
Madhani lab S. pombe 43K euchromatin tiling array |
| GPL9820 |
Madhani lab S. pombe 41K heterochromatin tiling array |
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| Samples (26)
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| Relations |
| BioProject |
PRJNA122381 |
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