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SUMMARY:Tackling Topology with TopoStats - Dr Alice Pyne\, University of S
 heffield
DTSTART:20230517T133000Z
DTEND:20230517T143000Z
UID:TALK193819@talks.cam.ac.uk
CONTACT:Lisa Masters
DESCRIPTION:Nearly all processes that act on DNA alter its topology\, prod
 ucing knotted\, catenated\, and supercoiled forms. Determining how these v
 ariations in DNA topology affect fundamental DNA interactions is challengi
 ng because of the length scale at which they occur\, 100x less than the wa
 velength of light. High-resolution atomic force microscopy (AFM) is unique
  in its ability to visualise DNA structure and interactions in liquid with
  sub-molecular resolution without the need for labelling or averaging. Tec
 hnological developments in high resolution AFM now allow it to visualise s
 ingle DNA molecules in liquid with sub-molecular resolution measuring the 
 twist\, writhe\, and topology of individual molecules in liquid as they 
 ‘explore’ their complex conformational space. \n\nHowever\, a rate-lim
 iting step for the widespread adoption of AFM to solve problems inaccessib
 le to the traditional tools of structural biology is a lack of open softwa
 re pipelines to analyse the increasing volumes of data produced. Automated
  analysis tools/software pipelines for AFM would reduce reliance on an exp
 erienced researcher\, minimise selection bias and facilitate the growth of
  AFM as a quantitative imaging technique. We have developed TopoStats (www
 .github.com/AFM-SPM/TopoStats)\, an open-source Python utility that loads 
 raw AFM data and handles data cleaning and processing through to identific
 ation and characterisation of individual DNA molecules [1].\n\nWe use Topo
 Stats to quantify the effect of supercoiling on DNA structure\, demonstrat
 ing that DNA under superhelical stress is far richer in structure\, e.g.\,
  containing kinks and defects\, than can be observed in short linear seque
 nces [2]. We have built on this foundation to develop tools that can accur
 ately identify\, isolate and trace the structure of individual DNA molecul
 es\, automatically pinpointing DNA crossings even in complex DNA structure
 s such as knots and catenanes. Our new image analysis routines can almost 
 unambiguously automatically identify under- and over-passing segments of D
 NA at each crossing\, thus allowing full identification of the knot/catena
 ne type and chirality. The information obtained within the AFM is much ric
 her than purely a measure of topology\, showing the heterogeneity in struc
 tures within each topological population. We propose that our combination 
 of high-resolution microscopy and automated analysis could enable the fiel
 d to probe how variations in the local structure and conformation of topol
 ogically complex DNA affect its interactions with essential cellular prote
 ins [3]. \n\nReferences:\n[1]	Beton\, JG et al. Methods 193\, 68-79 (2021)
  \n[2]	Pyne\, ALB*\, Noy A* et al. Nature Communications. 12\, 1053 (2021)
 \n[3]	Dos Santos\, A et al. Nature Communications (accepted)
LOCATION:Unilever Lecture Theatre\, Yusuf Hamied Department of Chemistry
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