The regulatory role of decoy transcription factor binding sites
How a gene responds to a transcription factor (TF) can depend on the number of binding sites within that TF's target promoter. But what if there are additional binding sites somewhere else in the genome? Using both experiments and models we demonstrate that these sites can qualitatively change the dose-response of the gene in a manner that depends quantitatively on the number of sites. Many organisms' genomes contain large regions of repetitive DNA sequence of variable length. A less appreciated consequence of this variable length may be an indirect impact on gene expression if these repetitive regions contain TF binding sites.
HFSP Young Investigator Grant holder Narendra Maheshri and colleaguesauthored on Thu, 26 April 2012
Tandem repeats are repetitive DNA sequences whose length is highly variable in the genomes of individuals within a population. Variability in the repeat numbers arises because of recombination and strand-slippage during replication (Gemayel et al, 2010). Our HFSP Young Investigator Grant team set out to understand the role of these tandem repeat regions in modifying gene regulation. The team has already shown that changing numbers of tandem repeats within promoters could both fine-tune and dramatically alter gene expression (Vinces et al, 2009). This recent study focuses on the impact tandem repeats located far away (in genomic distance) from a gene could have on its expression (Lee & Maheshri, 2012).
We hypothesized that if the tandem repeat unit contained a transcription factor binding site, then multiple TF binding sites could serve as a competitive inhibitor, or decoy, and sequester a TF and alter gene expression at that TF’s target genes. This is the basis of decoy therapies, where many oligonucleotides containing TF binding sites are introduced into cells to alter gene expression. In fact, because TFs are being constantly synthesized and degraded in the cell, using a kinetic model one can show that even this simple outcome will not occur if the decoy-bound TF is protected from degradation, but the unbound TF is not. We did find that repeated decoy sites to a transcriptional activator reduced expression of targeted genes, suggesting that DNA-bound TFs are degraded at the same rate as unbound TFs.
In the absence of decoys, the dose-response between the activator and gene expression was linear, and then eventually saturated. When the decoys were added, not only did gene expression decrease at any given dose of activator, but the shape of the dose-response curve changed from a linear to a steeper, threshold-like response. When analyzed in the context of a kinetic model, the implication is that the binding of the activator to the decoy sites was 10-100X stronger than binding to the promoter. This was unexpected because the decoy sites are identical in sequence to those in the promoter. Using chromatin immunoprecipiation, we confirmed that the activator was much more likely to bind the decoy sites versus the promoter when the repeated decoy region was less occupied. When the repeated decoy region was more occupied, the activator no longer bound the remaining unoccupied decoy sites as strongly. The qualitative trend occurred regardless even when we changed whether the repeated region was in different regions of the genome or in a plasmid.
Our findings were that: 1) repeated decoy sites located far from a promoter could affect gene expression by sequestering a TF, 2) TF binding to a promoter binding site in a context that could potentially activate gene expression is somehow weaker than binding to the same site outside a promoter context and 3) there may be a negative cooperativity in binding of a TF to repeated DNA binding sites that are clustered. The unexpected differences in TF binding strength can lead to qualitative changes in the behavior of gene regulatory networks in which target genes are embedded, a potential molecular link to explain how changes in the number of tandem repeats containing TF binding sites change an organism’s phenotype.
A regulatory role for repeated decoy transcription factor binding sites in target gene expression. Lee TH and Maheshri N. Mol Syst Biol. (2012) 8:576
Variable tandem repeats accelerate evolution of coding and regulatory sequences. Gemayel R, Vinces MD, Legendre M and Verstrepen KJ. Annu Rev Genet. (2010) 44:445-477.
Unstable tandem repeats in promoters confer transcriptional evolvability. Vinces MD, Legendre M, Caldara M, Hagihara M and Verstrepen KJ. Science. (2009) 324(5931):1213-1216.