Poster abstracts

Poster number 24 submitted by Ran Furman

ppGpp and DksA; global regulators of stress response

Ran Furman (Department of Microbiology, the Ohio State University), Georgiy A. Belogurov (Department of Microbiology, the Ohio State University), Irina Artsimovitch (Department of Microbiology, the Ohio State University)

Abstract:
Guanosine tetraphosphate (ppGpp) is one of the major regulators of stress response in bacteria. Numerous in vivo studies demonstrated large ppGpp effects on transcription, however, ppGpp alone displayed only minor effects in a purified in vitro system. To explain this discrepancy, the dnaK suppressor protein (DksA), was suggested to potentiate ppGpp effect in the cell. Indeed, DksA was shown to stimulate ppGpp activity in vitro. In this work, we performed a kinetic analysis of ppGpp and DksA effect at λPR promoter that provided several new insights:
A.ppGpp alone has a low affinity to the E. coli RNA polymerase (RNAP);
B.DksA inhibits transcription at the λPR promoter;
C.ppGpp increases DksA affinity to RNAP by more than 30-fold.
This analysis, together with the observations that, in contrast to ppGpp
whose levels rise dramatically during starvation, DksA is constitutively
expressed in the cell, led us to the following model:
Upon stress, ppGpp levels increase in the cell and consequently the
affinity of DksA to RNAP which leads to pleiotropic effects on
transcription. Interestingly different substitutions in DksA that increase
its affinity to RNAP may in part bypass the requirement for ppGpp.

References:
Murphy H. and Cashel M. Methods Enzymol. (2003), 371:596-601.
Paul B.J. et al., Cell. (2004),118(3):311-22.
Perederina A. et al., Cell. (2004), 118(3):297-309.
Rutherford S.T. et al., Genes Dev. (2009), 23(2):143-6.
Artsimovitch I. et al., Cell. (2004), 117(3):299-310
Vassylyeva M.N. et al., Acta Crystallogr D Biol Crystallogr. (2004),60:1611-3.
Vrentas C.E. et al., J Mol Biol.( 2008), 377(2):551-64
Sato M. et Al., Chembiochem. (2009), 10(7):1227-33.
Magnuson L.U. et al., J Bacteriol. (2007),189(14):5193-202.
Paul B.J. et al., Proc Natl Acad Sci. (2005),102(22):7823-8
Rutherford S.T. et al., Genes Dev. (2009), 23(2):236-48.
Charles E. et al., Nature (1995), 373, 636 – 640
Lamour V. et al., J Mol Biol. (2006),356(1):179-88
Blankschien M.D. et al., EMBO (2009), on line publication
Laptenko O. et al., EMBO J. (2006),25(10):2131-41.

Keywords: DksA, ppGpp, RNAP