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I am studying a particular gene whose expression decreases in certain types of cancer. I'm interested in finding out the mechanisms for this downregulation, specifically, which transcription factors and signalling pathways are involved. I have a hypothesis for the pathway involved, but in case that hypothesis turns out to be incorrect, I want to look into some ways of approaching the problem in an unbiased manner. Here are my thoughts for some starting approaches:
1. Feed the promoter DNA for this gene into a computer program to look for transcription factor binding sites. (https://abc.med.cornell.edu/education/introtobio/t-promoter.html seems to list a number of resources. Does anyone have experience with any of these?) However, a lot of gene regulation in mammals occurs though distal enhancer sequences. Are there computational or experimental ways to identify enhancers for a target gene?
2. Generate a reporter cell line by placing GFP under the control of my target gene's promoter and perform an siRNA screen to see what perturbations increase/decrease expression of the reporter.
Are there better ways of computationally and/or experimentally approaching this problem? Any suggestions or pointers to helpful references would be appreciated.
1. Feed the promoter DNA for this gene into a computer program to look for transcription factor binding sites. (https://abc.med.cornell.edu/education/introtobio/t-promoter.html seems to list a number of resources. Does anyone have experience with any of these?) However, a lot of gene regulation in mammals occurs though distal enhancer sequences. Are there computational or experimental ways to identify enhancers for a target gene?
2. Generate a reporter cell line by placing GFP under the control of my target gene's promoter and perform an siRNA screen to see what perturbations increase/decrease expression of the reporter.
Are there better ways of computationally and/or experimentally approaching this problem? Any suggestions or pointers to helpful references would be appreciated.
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