The School of Molecular and Cellular Biology at the University of Illinois at Urbana-Champaign

Our work focuses on the genome caretakers involved in homologous recombination and nucleotide excision repair. The following systems are being currently pursued in my lab:

Motor components of the DNA repair machineries, in particular DNA helicases XPD, Bach1, Fbh1. Central to many DNA-repair machineries are DNA helicases, ubiquitous molecular motors that use the energy of nucleoside triphosphate (NTP) hydrolysis to move directionally along nucleic acids. Translocation by helicases is coupled to other thermodynamically unfavorable processes including separation of nucleic acid duplexes and disassembly of protein-nucleic acid complexes. The question we try to answer is how these structurally similar enzymes perform an amazingly diverse set of activities, how they utilize unique structural features incorporated into otherwise conserved motor cores and how other players in the genome maintenance pathways modulate activities of selected helicases adapting them to desired cellular tasks.

Recombination mediators: Lesions affecting both strands of the DNA duplex are constantly generated in the cell and have to be mended to prevent genomic instability leading to cancer or cellular senescence. Most accurately this is achieved through the process of homologous genetic recombination, which depends on assembly of the Rad51 recombinase into continuous nucleoprotein filament on ssDNA generated at the site of damage. Rad51 nucleoprotein filament formation is opposed by ssDNA sequestering by ssDNA binding proteins and is highly controlled by a group of recombination mediator proteins. We study one of these recombination mediators, Rad52 protein and its effect of Rad51 activities.

APPROACHES:

We study the DNA repair processes at the most fundamental level by first deconstructing the macromolecular ensembles orchestrating the DNA repair events down to the level of individual proteins. By combining classical and single-molecule biochemistry for analysis of the key players in these DNA repair pathways, we then investigate their molecular mechanism and how other protein partners affect their action. These studies allow us to gain fundamental biochemical knowledge of accurate DNA repair and help us to identify features of the genome caretaker proteins that can be exploited in designing the new therapeutics.

Representative Publications

Spies M, Ha T., Inching over hurdles: how DNA helicases move on crowded lattices. (2010) Cell Cycle 9(9)[1742-9 PMID: 20436294

Pugh RA, Honda M, Spies M., Ensemble and single-molecule fluorescence-based assays to monitor DNA binding, translocation, and unwinding by iron-sulfur cluster containing helicases.(2010) METHODS 51(3) 313-21 PMID: 20167274

Grimme JM, Honda M, Wright R, Okuno Y, Rothenberg E, Mazin AV, Ha T, Spies M., Human Rad52 binds and wraps single-stranded DNA and mediates annealing via two hRad52-ssDNA complexes (2010) NAR 38(9)2917-30 PMID: 20081207; PMCID: PMC2875008

Honda, M., Park, J., Pugh, R. A., Ha, T., and Spies, M., Single-molecule analysis reveals differential effect of ssDNA binding proteins on DNA translocation by XPD helicase: (2009) Mol.Cell 35(5) 694-703 PMID: 19748362; PMCID: PMC2776038

Rothenberg, E., Grimme, J. M., Spies, M. and Ha, T.,Rad52 protein mediates directionally biased homology search and DNA annealing through continuous association of two Rad52-ssDNA complexes. (2008) PNAS 105 (51) 20274-20279; PMID: 19074292; PMCID: PMC2629295

Pugh, R. A., Lin, Y., Eller C., Leesley H., Cann, I. K.O., and Spies, M. Ferroplasma acidarmanus RPA2 facilitates efficient unwinding of forked DNA substrates by monomers of XPD helicase. (2008) JMB 383(5): 982-98; PMID: 18801373

Lin, Y., Lin, L.-J., Sriratana, P., Coleman, K., Ha, T., Spies, M., Cann, I.K.O. Engineering of functional replication protein A homologs based on insights in evolution 1 of 2 oligonucleotide/oligosaccharide binding (OB) folds. (2008); J. Bac, 190 (17) 5766-5780

Pugh, R.A., Honda, M., Leesley, H., Thomas, A., Lin, Y., Nilges, M.J., Cann, I.K.O., Spies, M. The iron-containing domain is essential in RAD3 helicases for coupling of ATP hydrolysis to DNA translocation and for targeting the helicase to the ssDNA-dsDNA junction. (2008) JBC, 283(3):1732-43.

Spies, M., Amitani, I., Baskin, R.J., and Kowalczykowski, S. C. Translocation by single-molecules of a RecBCD motor mutant reveals a switch in motor subunit usage at χ. (2007) Cell, 131: 694-705;

Grimme, J. M., and Spies, M., "DNA helicases in homologous recombination" (in "Molecilar Genetics, Biophysics and Medicine Today" (Bresler Memorial Lectures –II) Editor: Vladislav Lanzov), PNPI Press, 2007, St. Petersburg, Russia ISBN 5-86763-197-4) 123-140

Complete Publications List