kill-frap.tex

\documentclass[a4paper,oneside,onecolumn,article,draft]{memoir}
\counterwithout{section}{chapter}

\input{preamble}

\author{David Miguel Susano Pinto \and Kevin F. Sullivan \and Andrew Flaus}
\title{Application of FRAP to histones in human cell nuclei}

\begin{document}

  \maketitle

  \begin{abstract}
    Nucleosomes enable the stable compaction of almost all eukaryotic
    genomes but also require dynamic properties to enable access to
    the packaged DNA sequences.  The stability of core histones within the
    nucleosome should be reflected in their capability for dynamic exchange by
    Fluorescence Recovery After Photobleaching (FRAP).  To assay the
    effect of histone SWI/SNF INdependence (SIN)
    mutants known to destabilise nucleosomes
    \textit{in vitro} and in \species{S. cerevisiae}, we sought to
    apply FRAP to chromatin in mammalian cell lines.  This uncovered a
    number of challenges resulting from cell motility, nuclear movement
    within the cell, and chromatin motion within the nucleus with the
    long time frames required for FRAP of histones.  We were able to
    compensate for the former difficulties by a combination of cell
    biological and computational techniques, but we were unable to
    establish an appropriate approach to compensate for motion of the immobile
    binding sites required for standard FRAP analysis.
    Visualisation using photoactivated tagged histones in inverse FRAP
    demonstrated the extent of this chromatin motion
    and a further complexity arising from complex
    non-homogenous channelling tagged histone
    during diffusion.  This reveals the
    limitations of FRAP over extremely long time scales, and suggests
    that this technique is unsuitable for quantitative measurement of
    histone dynamics in the mammalian nucleus.
  \end{abstract}

  \input{sections/intro}
  \input{sections/methods}
  \input{sections/results}
  \input{sections/discussion}
  \bibliography{references}

\end{document}