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Studying transcriptional interactions in single cells at sufficient resolution
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
2011 (English)In: Current Opinion in Biotechnology, ISSN 0958-1669, E-ISSN 1879-0429, Vol. 22, no 1, 81-86 p.Article, review/survey (Refereed) Published
Abstract [en]

Our ability to dissect and understand the principles of gene regulatory circuits is partly limited by the resolution of our experimental assays. In this brief review, we discuss aspects of gene expression in microbial organisms apparent only when increasing the experimental resolution from populations to single cells and sub-cellular structures, from snap-shots to high-speed time-lapse movies and from molecular ensembles to single molecules.

Place, publisher, year, edition, pages
2011. Vol. 22, no 1, 81-86 p.
National Category
Biological Sciences
Identifiers
URN: urn:nbn:se:uu:diva-149731DOI: 10.1016/j.copbio.2010.10.004ISI: 000287840700012PubMedID: 21071200OAI: oai:DiVA.org:uu-149731DiVA: diva2:405350
Available from: 2011-03-22 Created: 2011-03-22 Last updated: 2017-12-11Bibliographically approved
In thesis
1. How precise is cyclic life?: Insights during a single molecule revolution of the bacterial cell cycle.
Open this publication in new window or tab >>How precise is cyclic life?: Insights during a single molecule revolution of the bacterial cell cycle.
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Bacterial cells reproduce by doubling in size and dividing. The molecular control systems which regulate the cell cycle must do so in a manner which maintains a similar cell size over many generations. A cell can under conditions of fast growth conclude cell cycles in shorter time than the time required to replicate its chromosome. Under such conditions several rounds of replication are maintained in parallel and a cell will inherit replication processes which were initiated by an ancestor. To accomplish this the cell has to initiate and terminate one round of replication during each cell cycle.

To investigate the effects of the cell cycle on gene-regulation in the gut bacterium Escherichia coli, an experimental method combining microfluidics, single molecule fluorescence microscopy and automated analysis capable of acquiring an arbitrary number of complete cell cycles per experiment was developed. The method allowed for the rapid exchange of the chemical environment surrounding the cells. Using this method it was possible to measure the dissociation time of the transcription factor molecule, LacI-Venus, from the native lactose operator sequence, lacO1, and an artificially strong operator, lacOsym, in vivo. The results indicated that regulation of gene-expression from the lactose operon does not occur at equilibrium in living cells. Furthermore, by studying the intracellular location of non-specifically interacting transcription factor molecules it was possible to determine that these do not form long-lived gradients inside the cell as was previously proposed.

By studying the replication machinery and the origin of replication it was found that replication is initiated according to a cell volume per origin which did not vary over different growth conditions. Further, division timing was found to be determined by the initiation event to occur after a fixed time-delay. A consequence of this mode of regulation is an uncertainty relation between the size at birth and the cell cycle time, in which cells will vary more in in the cycle time during conditions of slow growth as compared to fast growth and vary more in birth length during conditions of fast growth as compared to slow growth.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2014. 77 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1204
Keyword
E.coli, cell cycle, replication initiation, transcription factor, gene-regulation, single molecule microscopy, microfluidics
National Category
Biochemistry and Molecular Biology
Research subject
Biology with specialization in Molecular Biotechnology
Identifiers
urn:nbn:se:uu:diva-235546 (URN)978-91-554-9104-8 (ISBN)
Public defence
2014-12-05, BMC A1:111a, Husarg. 3, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2014-11-13 Created: 2014-11-05 Last updated: 2015-02-03

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Walldén, MatsElf, Johan

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