Question
Present evidence for the hypothesis that an insulator blocks enhancement by interacting with nearby enhancers and promoters. What are the difficulties in generalizing this hypothesis to all insulators?
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Insulators are DNA elements that can prevent the interaction between enhancers and promoters, thereby blocking the enhancement of gene expression. Show more…
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When enhancers were initially found to influence transcription from many thousands of nucleotide pairs away from the promoters they control, two principal models were invoked to explain this action at a distance. In the "DNA looping" model, direct interactions between proteins bound at enhancers and promoters were proposed to stimulate transcription initiation. In the "scanning" or "entry-site" model, RNA polymerase (or another component of the transcription machinery) was proposed to bind at the enhancer and then scan along the DNA until it reached the promoter. These two models were tested using an enhancer on one piece of DNA and a $\beta$ -globin gene and promoter on a separate piece of DNA (Figure $\mathrm{Q} 8-9$ ). The $\beta$ -globin gene was not expressed when these two separate pieces of DNA were introduced together. However, when the two segments of DNA were joined via a linker (made of a protein that binds to a small molecule called biotin), the $\beta$ -globin gene was expressed. Does this experiment distinguish between the DNA looping model and the scanning model? Explain your answer.
A graduate student came up with the following idea for identifying insulators in the Drosophila genome: Perform an experiment like the one described in Problem $12,$ but instead of using an enhancerless construct, use one that contains an enhancer, and screen for lines that do not express GFP. a. What is wrong with this experimental design? b. Can you think of a different experiment that you could perform to identify insulators?
If you insert a $\beta$ -galactosidase gene lacking its own transcription control region into a cluster of piRNA genes in Drosophila , you find that $\beta$ -galactosidase expression from a normal copy elsewhere in the genome is strongly inhibited in the fly's germ cells. If the inactive $\beta$ -galactosidase gene is inserted outside the piRNA gene cluster, the normal gene is properly expressed. What do you suppose is the basis for this observation? How would you test your hypothesis?
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