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Real-time observation of Cas9 Activity


Binding specificity of Cas9–guide RNA complexes to DNA is important for genome-engineering applications; however, how mismatches influence target recognition/rejection kinetics is not well understood. Here, the Ha group, in collaboration with the Doudna lab at UC Berkeley, used single-molecule FRET to probe real-time interactions between Cas9–RNA and DNA targets. The bimolecular association rate is only weakly dependent on sequence; however, the dissociation rate greatly increases upon introduction of mismatches proximal to protospacer-adjacent motif (PAM), demonstrating that mismatches encountered early during heteroduplex formation induce rapid rejection of off-target DNA. In contrast, PAM-distal mismatches up to 11 base pairs in length, which prevent DNA cleavage, still allow formation of a stable complex, suggesting that extremely slow rejection could sequester Cas9–RNA, increasing the Cas9 expression level necessary for genome-editing, thereby aggravating off-target effects. The team also observed at least two different bound FRET states that may represent distinct steps in target search and proofreading. Refer to:

1/21/2014 Siv Schwink

Professor Karin Dahmen has been elected a Fellow of the American Physical Society “for establishment and exploring the deep connections between non-equilibrium phase transitions and avalanche phenomena in diverse fields encompassing materials, geophysics and neuroscience."

Dahmen is a condensed matter theorist whose work in systems theory bridges condensed matter physics, mathematical physics, geophysics, biophysics, and materials science. Dahmen analyzes and models the statistical properties of both living and non-living systems to uncover the universality and predictability of complex behavior patterns within widely varied systems—from neuron-firing avalanches in the brain, to the behavior of electrons in high-temperature superconductors; from the effect of uneven distribution of nutrients on bacterial population dynamics, to equilibrium and nonequilibrium-critical behavior in magnetic systems.

Department Head and Professor Dale Van Harlingen shares, “It is great to see Karin get this well-deserved recognition.  Her pioneering work on modeling the dynamics of complex systems is finding applications in many fields ranging from vortices in strongly-correlated condensed matter systems, to population dynamics in biological systems, and to avalanches and earthquakes critical to the environment. Her ability to extract unifying mathematical patterns from highly complex data sets has established her as a leading theorist working in systems theory.”

Since 2006, Dahmen’s group has forged several new directions for interdisciplinary systems theory research. Whether working at the macro- or nanoscale, Dahmen’s group applies a broad range of ideas—from statistical physics, dynamical systems theory, disordered-systems theory, the theory of phase transitions, the renormalization group, hydrodynamics, computational physics, and mathematical physics—to develop new analytical methods and tools. Dahmen’s team proposed a unifying theory of slip avalanches; introduced new methods for analyzing strongly correlated electronic systems to answer persistent questions about complex electronic patterns at the nanoscale on the surface of high-temperature superconductors; and, in collaboration with experimentalists at of the Center for the Physics of Living Cells (CPLC), developed data analysis tools to extract signals from noisy fluorescence resonance energy transfer  (FRET) data, using hidden Markov modeling techniques.

Dahmen’s theoretical work has implications for many real-world applications, including extracting the microstructure and lifetime of materials, mapping the functional network structure in brain regions, developing nondestructive materials testing and failure prediction, and hazard prediction and prevention ranging from large earthquakes to individual epileptic seizures.

At the University of Illinois at Urbana-Champaign, Dahmen is a member of both the Institute for Condensed Matter Theory and of the Frederick Seitz Materials Research Center’s Materials Computation Center. She is also a co-principal investigator at the Center for the Physics of Living Cells, an NSF Physics Frontiers Center, and an affiliate at the NASA-funded Institute for Genomic Biology.

Dahmen is the recipient of several honors. She received the 2011 Xerox Senior Faculty Research Award, was named a Beckman Fellow at the U. of I. Center for Advanced Study in 2001, and was selected an Alfred P. Sloan Research Fellow, also in 2001.

Dahmen received her Vordiplom in physics from the Universität Bonn, Germany, in 1989, and her master’s and doctoral degrees in physics from Cornell in 1994 and 1995, respectively. Before joining the faculty at Illinois in 1999, and completed postdoctoral work as a Junior Fellow at Harvard University.