title: Inferring diffusion dynamics from FCS in heterogeneous nuclear environments.
abstract: Fluorescence correlation spectroscopy (FCS) is a noninvasive technique that probes the diffusion dynamics of proteins down to single-molecule sensitivity in living cells. Critical mechanistic insight is often drawn from FCS experiments by fitting the resulting time-intensity correlation function, G(t), to known diffusion models. When simple models fail, the complex diffusion dynamics of proteins within heterogeneous cellular environments can be fit to anomalous diffusion models with adjustable anomalous exponents. Here, we take a different approach. We use the maximum entropy method to show-first using synthetic data-that a model for proteins diffusing while stochastically binding/unbinding to various affinity sites in living cells gives rise to a G(t) that could otherwise be equally well fit using anomalous diffusion models. We explain the mechanistic insight derived from our method. In particular, using real FCS data, we describe how the effects of cell crowding and binding to affinity sites manifest themselves in the behavior of G(t). Our focus is on the diffusive behavior of an engineered protein in 1) the heterochromatin region of the cell's nucleus as well as 2) in the cell's cytoplasm and 3) in solution. The protein consists of the basic region-leucine zipper (BZip) domain of the CCAAT/enhancer-binding protein (C/EBP) fused to fluorescent proteins.
authors.affiliation: Department of Physics, IUPUI, Indianapolis Indiana.
authors.forename: Konstantinos
authors.initials: K
authors.lastname: Tsekouras
authors.affiliation: Integrated Nanosystems Development Institute, IUPUI, Indianapolis Indiana.
authors.forename: Amanda P
authors.initials: AP
authors.lastname: Siegel
authors.affiliation: Department of Cellular & Integrative Physiology, Indiana University School of Medicine, Indianapolis, Indiana.
authors.forename: Richard N
authors.initials: RN
authors.lastname: Day
authors.affiliation: Department of Physics, IUPUI, Indianapolis Indiana; Department of Cellular & Integrative Physiology, Indiana University School of Medicine, Indianapolis, Indiana. Electronic address: stevenpresse@gmail.com.
authors.forename: Steve
authors.initials: S
authors.lastname: Pressé
mesh_terms: D000818:Animals; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000737:chemistry / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004058:Diffusion; D019277:Entropy; D006570:Heterochromatin / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods*; D008954:Models, Biological; D008958:Models, Molecular; D013269:Stochastic Processes
chemical_list: D022763:CCAAT-Enhancer-Binding Protein-alpha; D006570:Heterochromatin