Force-Dependence of Lac-Repressor Mediated DNA Loop Formation

Abstract

Protein-mediated DNA looping is a ubiquitous motif in transcriptional control schemes. Formation of these loops is driven by thermal fluctuations of the substrate DNA, which in turn are known to be exquisitely sensitive to mechanical constraints on the DNA. Because DNA in vivo is subject to a complex micromechanical environment, it is intriguing to study the effect of mechanical tension in the substrate DNA on the formation of these regulatory loops to investigate the role of mechanics in controlling gene regulation. For this purpose, we measured the formation and breakdown rates of lac repressor-mediated DNA loops under tension using constant-force axial optical tweezers.We observed that an incremental force of less than 100 femtonewtons is sufficient to reduce loop formation rate about sevenfold in a construct with an inter-operator spacing of 305 bp. This result suggests the possibility of mechanical pathways to control gene expression with forces that are two orders of magnitude lower than other typical intracellular forces acting on DNA, such as the forces exerted by RNA polymerase and molecular motors. Moreover, we developed a model that quantifies the relation between the force sensitivity of the loop formation rate and the angle between the incoming and outgoing DNA strand in the loop as a way to infer loop topology from our micromechanical measurements. We conclude that the LacI-mediated DNA loop prefers an anti-parallel loop topology over a parallel conformation.