Multiple Lac-Mediated Loops Revealed by Bayesian Statistics and Tethered Particle Motion

Abstract

The bacterial transcription factor LacI loops DNA by binding to two distinct binding sites on DNA. Despite being one of the most well-studied model systems for transcriptional regulation, the number of possible loops, and their structures, are unclear. To clarify this question, we have developed a new analysis method for tethered particle motion (TPM), a versatile single-molecule technique commonly used to study Lac-mediated looping and other DNA-protein interactions in vitro. The method, called vbTPM, is based on a variational Bayes treatment of hidden Markov models. It learns the number of distinct states (e.g., DNA-protein conformations) in a TPM trajectory directly from the data with better resolution than existing methods, and also corrects for common experimental artifacts. Studying short (about 100 bp) LacI-mediated loops, we are able to resolve states and interconversion patterns that are best explained in terms of at least three distinct loop structures. These results indicate that changes in Lac conformation and DNA binding orientation both contribute to the repertoire of LacI-mediated loops, and provides new input for quantitative models of looping and transcriptional regulation.

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