On-Chip Expression of Transcription Factors and Force-Based Interaction Analysis with DNA

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Proteins regulate gene expression and their products may in turn regulate other genes. A highly en-tangled multi-component network arises upon which life is built. Needless to mention that deciphering this network requires the tracing of an exceedingly growing number of interaction partners and their quantification in order to characterize the quality of their interactions. A variety of highly parallel assays were developed in the past and most of them were hampered by a lack of sufficient sensitivity and/or selectivity. More independent measured values of a given interaction are needed to expand the parameter space into which these interactions are projected. The proposal here suggests to measure the interaction forces between proteins and DNA and by means of this to provide an additional parameter that characterizes the interaction. A highly parallel chip format is proposed which combines the expression of the proteins in soft rubber micro-fluidics with fluorescence-based binding force measurements via molecular force balances. As proof of principle a set of transcription factors from yeast will be tested against a matrix of ten different target sequences and for six different unbinding forces and compared to literature results. An expansion by at least one order of magnitude in integration density is envisioned which then allows the quantitative screening of a large number of interaction partners in particularly also weak binders in parallel. We will then expand our collaborations to characterize drosophila transcription factors based on interaction forces.