Building Pathway Sets
PATHWAY PROFILING PLATES
In order to build a pathway profiling panel, we indentified a set of promoters that are often regulated as part of the same signaling pathway yet tend to vary in response to different stimuli. A library of elements that respond similarly to every stimulus would not yield useful profiles for comparing compounds or conditions. We first selected promoters from an initial candidate set that showed a significant response (either induction or repression) under the tested conditions. However, we were also interested in including other elements that showed either no response or a subtle response in our initial testing. While these elements might not seem interesting based on a single experiment, other characteristics highlight that they are likely to be relevant for studies in other cell lines or with other treatments: 1) known target from literature; 2) alternative promoter for a known target or strong responder; 3) strong transcription factor binding motif scores; 4) transcription factor binding data from ChIP-type experiments.
BIOMARKER SETS
The goal for building a set of key responders is somewhat different than building a profiling panel since we are looking for promoters that may serve as consistent “biomarkers” for activation of a pathway in a variety of settings. The best promoter to use as a biomarker may vary depending on context, so we have selected a small set of robust responders from each pathway. Researchers may then identify their ideal markers by surveying activity across this small set in their experimental systems. We have also included two housekeeping or random promoters that did not respond to pathway activation in initial testing that may be used to correct for non-specific changes in overall signal based on a stimulus.