Tuesday 02/04/2014

11:00a – 8:00p, 9:30p – 10:30p

Ph project

Manuscript writing

  • working on model section
    • finished description of first model (on-rate or off-rate limited)
    • finished writing description of second model, ~3:30p, (chromatin availability limited)
  • working on methods section
    • wrote description of STORM imaging
    • wrote description of antibody labeling
    • wrote description of image analysis and clustering
    • wrote description of modeling / polymer-simulations
  • working on discussion
    • Basic discussion of clusters needs overhaul.


  • TO DO: Redesigning Figure 5 (model figure).
    • change kinetic panels to first set.
    • remove images (too many panels)
    • redo plots in second set of models


  • finished reading Tijan Lab Elife article on fastFISH.
    • Nice kinetic measurements of RNAP binding, RNA appearance, RNAP release and RNA release.
    • All with T7 on glass substrate, would be nice to eukaryotic polymerases (in vivo?) Probably requires a lot more signal and doesn’t require the ultra fast hybridization since T7 is blazing fast (binding 1 per second, elongating 300 bp/s or something like that)
  • Pivot algorithm improvements
  • Beautiful review on regulation of gene expression in the genomic context by Taylor Atkinson and Marc Halfon: http://journals.sfu.ca/rncsb/index.php/csbj/article/view/csbj.201401001
  • Simon and Kingston: Occupying chromatin: Polycomb mechanisms for getting to genomic targets, stopping transcriptional traffic, and staying put.
    • ‘central mechanistic question, how PcG complexes shut down transcription, has been a tough nut to crack’. (i.e. not yet worked out?)
    • long standing idea of altered template accessibility called into question (no new citations here though, I suspect they follow in a more detailed discussion after the intro).
    • PcG targeting in mammals more RNA directed (e.g. Xist, HOTAIR). Though deletion of HOTAIR + flanking regions has no phenotype.
    • no direct evidence yet, merely correlative (knock this down, change H3K27me3). Want to see, mutate this binding site which affects interaction, change repression. Observe lncRNA directly at the site where the change (PcG binding) is happening.
    • emerging evidence challenges the central role of K27me3 for targeting PRC1.
    • Yuan et al 2012 study implies K27me3 accumulates after onset of silencing (sure, but PcG dependent silencing?)
    • perturb K3K27me3 levels not dramatically disturb immunostaining of PRC1 (presumably in polytene).
    • Pc and Ph unnecessary of ubiquitination activity of PRC1, and they compete for a position in the complex with KDM2, an enhancer of ubiquitination. (genetically though they are key elements of PcG silencing).

(8:00p-9:30p. Replace broken bike chain. Attempt and failed to replace worn out rear gear cassette)

Chromatin Project


  • STORM2 available again
  • Finish imaging current recently stained cells (G05 and G09).
  • Check age of fixed cells, repeat attempt of double stains
  • Double stains done so far (none of which I’ve found time to analyze because of the Ph project take-over).
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