CRISPR-Cas9 Genome Interrogation: A Facilitated Subdiffusive Target Search Strategy

The functional application of RNA-guided CRISPR-associated Cas9 protein, a bacterial immune system-based protein complex, via which in vivo, highly specific, and well-regulated, gene-editing processes are being monitored at an unprecedented level, has led to remarkable progress in genetic engineering and technology. The complicated in vivo process of genome interrogation followed by gene editing by the Cas9 complex was recently reported by Knight et al. (Science, 2015, 350, 823-826) using an elegant single-particle tracking method, aided by the two-photon fluorescence correlation spectroscopic technique. In contrast to the usually observed fast target-searching and protein-binding events in biophysical systems, an interesting slow genome-interrogation process by the RNA-guided CRISPR-Cas9 system through a crowded chromatin environment of a mammalian cell has been revealed in Knight et al.'s study. Motivated by this experiment, in this paper, we provide a generalized theoretical framework to capture this particular target-searching mechanism of the CRISPR-Cas9 protein complex. We show that an analysis on the basis of 3D subdiffusion under a cylindrical volume, created by several nonspecific off-target interactions from the DNA strands, can capture the essential details of the process. Moreover, on the basis of this model, we quantify the dynamics of this process and estimate the survival probability, first passage time, and the intensity correlation function of the tagged proteins of the experiment. The results from our theoretical predictions are found to be consistent with the experimental observations, and hence, seem to provide a plausible microscopic picture of the process.