Single-Cell Biophysics: Measurement, Modulation, and Modeling

Single-Cell Biophysics: Measurement, Modulation, and Modeling

Tuesday Speaker Abstracts

Single-Molecule Investigations of DNA Replication and Repair in Living Bacteria

Julie Biteen

University of Michigan, Ann Arbor, MI, USA.

By beating the diffraction limit that restricts traditional light microscopy, single-molecule

fluorescence imaging is a precise, non-invasive way to sensitively probe position and dynamics.

Our lab has been developing new methods to locate, track, and analyze single molecules to

measure structure, dynamics, and cooperativity in live bacterial cells. I will discuss how we are

measuring and understanding the dynamical interactions essential for DNA mismatch

recognition and DNA replication in living

Bacillus subtilis

cells. In particular, we have

integrated our nanoscopic biophysical tools with biochemical and genomic approaches to

understand how the mismatch repair protein MutS efficiently identifies DNA mismatches during

real time in living cells, to determine that the replicative polymerase PolC is continuously

recruited to and released from a centrally located replisome, and to measure how the various

components of the multi-protein replication machinery are organized in bacterial cells. Overall,

our results provide a mechanistic model for the dynamical nature of these fundamental processes

in live cells.

Single-Molecule Visualization of Bacterial DNA Repair in Live Cells

Antoine M. van Oijen

University of Wollongong, Wollongong, NSW, Australia.

We are using single-molecule fluorescence methods to visualize the behaviour of individual,

fluorescently labelled molecules inside living cells. Using these approaches, we aim to

understand the relationship between DNA replication and repair. In particular, we study aspects

related to the bacterial SOS damage response: how does the RecA protein coat single-stranded

DNA after DNA damage stalls replication and how do the subsequently upregulated translesion

synthesis DNA polymerases gain access to the lesions in the DNA? Further, we investigate the

dynamics of transcription-coupled repair as mediated by the E. coli Mfd protein. These studies

not only lead to new insights in molecular pathways, they also provide information on the critical

role played by spatial and temporal regulatory mechanisms.