Abstract
Characterizing how genomic sequence interacts with trans-acting regulatory factors to implement a program of gene expression in eukaryotic organisms is critical to understanding genome function. One means by which patterns of gene expression are achieved is through the differential packaging of DNA into distinct types of chromatin. While chromatin state exerts a major influence on gene expression, the extent to which cis-acting DNA sequences contribute to the specification of chromatin state remains incompletely understood. To address this, we have used a fission yeast sequence element (L5), known to be sufficient to nucleate heterochromatin, to establish de novo heterochromatin domains in the Schizosaccharomyces pombe genome. The resulting heterochromatin domains were queried for the presence of H3K9 di-methylation and Swi6p, both hallmarks of heterochromatin, and for levels of gene expression. We describe a major effect of genomic sequences in determining the size and extent of such de novo heterochromatin domains. Heterochromatin spreading is antagonized by the presence of genes, in a manner that can occur independent of strength of transcription. Increasing the dosage of Swi6p results in increased heterochromatin proximal to the L5 element, but does not result in an expansion of the heterochromatin domain, suggesting that in this context genomic effects are dominant over trans effects. Finally, we show that the ratio of Swi6p to H3K9 di-methylation is sequence-dependent and correlates with the extent of gene repression. Taken together, these data demonstrate that the sequence content of a genomic region plays a significant role in shaping its response to encroaching heterochromatin and suggest a role of DNA sequence in specifying chromatin state.
Highlights
Correct patterns of gene expression are established by orchestrated interactions among cis-regulatory elements, transacting factors and the surrounding chromatin environment
Our study focuses on the transition between two opposing chromatin states: euchromatin, which generally correlates with gene expression, and heterochromatin, which is typically refractive to gene expression
The effect of L5-integration on the chromatin environment of sequences within the ura4 locus was characterized by quantifying H3K9me2 and Swi6p levels throughout the region using chromatin immunoprecipitation (ChIP)
Summary
Correct patterns of gene expression are established by orchestrated interactions among cis-regulatory elements, transacting factors and the surrounding chromatin environment. The magnitude of H3K9me enrichment decreases over the Pnmt1-his3+sequences and remains reduced over the distal portion of the spacer DNA (Figure 3) These data demonstrate that the insertion of genic sequences within the spacer DNA attenuates the spread of heterochromatin and further support the hypothesis that the presence of genes within the ura and spbc2f12.03 neighborhoods limits heterochromatin spreading. It is interesting, that the Pnmt1-his3+ construct, despite being more highly transcribed than spcc330.06+ (Figure S5A), does not exhibit complete barrier activity (Figure 3), suggesting that factors other than high levels of transcriptional activity are required for complete barrier activity
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
