Determining how cells sense and adapt to fluctuating O2 levels in their environment is a fundamental problem in biology. For many prokaryotic and eucaryotic organism, the sensing of oxygen levels is essential to ensure an adequate supply of energy as well as to avoid the toxic effects of oxygen. In recent years, it has been shown that many members of the FNR family of transcription factors that play a key role in the anaerobic lifestyle of a wide group of prokaryotes, function in oxygen sensing. Thus by deciphering how FNAR activity responds to oxygen availability, we will obtain fundamental information on a process that is critical to the growth and survival of facultative microbes. In addition, our studies should provide key insights into some general properties of oxygen sensing that can be applied to both eucaryotic and prokaryotic cells. In the best-studies case of Escherichia coli, FNR contains a [4Fe-4S]+2 cluster that is required for dimerization and site-specific DNA binding. This [4Fe-4S]+2 cluster is oxygen sensitive and its conversion to a [2Fe-21S]+2 decreases dimerization and DNA binding in vitro. To explain how FNR activity is regulated in vivo, we propose that FNR is largely active under anaerobic conditions because the [4Fe-4S]+2 cluster is stable whereas, in the presence of oxygen, we propose that FNR is largely inactive due to its conversion to a [2Fe-2S]+2 intermediate or possibly an apo-FNR form that lacks a cluster. To determine whether the [4Fe-4S]+2 cluster conversion is sufficient to explain how FNR is inactivated under aerobic conditions in vivo, we will define the pathway of FNR inactivation in vivo. To determine how the presence of the [4Fe-4S]+2 cluster increases FNR activity, we will test the hypothesis that the [4Fe-4S]+2 cluster is required to achieve a conformation that is competent for dimerization. To further define the role of FNR as a global regulator of transcription we will characterize the regions of FNR involved in transcription activation. Our studies should rovide insights into conserved regulatory strategies for sensing changes in oxygen tension by a wide variety of prokaryotes including several pathogenic organisms.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM045844-13
Application #
6636028
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Program Officer
Anderson, James J
Project Start
1991-04-01
Project End
2005-03-31
Budget Start
2003-04-01
Budget End
2005-03-31
Support Year
13
Fiscal Year
2003
Total Cost
$316,800
Indirect Cost
Name
University of Wisconsin Madison
Department
Biochemistry
Type
Schools of Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Beauchene, Nicole A; Mettert, Erin L; Moore, Laura J et al. (2017) O2 availability impacts iron homeostasis in Escherichia coli. Proc Natl Acad Sci U S A 114:12261-12266
Myers, Kevin S; Park, Dan M; Beauchene, Nicole A et al. (2015) Defining bacterial regulons using ChIP-seq. Methods 86:80-8
Beauchene, Nicole A; Myers, Kevin S; Chung, Dongjun et al. (2015) Impact of Anaerobiosis on Expression of the Iron-Responsive Fur and RyhB Regulons. MBio 6:e01947-15
Mettert, Erin L; Kiley, Patricia J (2015) Fe-S proteins that regulate gene expression. Biochim Biophys Acta 1853:1284-93
Tolla, Dean A; Kiley, Patricia J; Lomnitz, Jason G et al. (2015) Design principles of a conditional futile cycle exploited for regulation. Mol Biosyst 11:1841-9
Miller, Halie K; Kwuan, Laura; Schwiesow, Leah et al. (2014) IscR is essential for yersinia pseudotuberculosis type III secretion and virulence. PLoS Pathog 10:e1004194
Park, Dan M; Kiley, Patricia J (2014) The influence of repressor DNA binding site architecture on transcriptional control. MBio 5:e01684-14
Mettert, Erin L; Kiley, Patricia J (2014) Coordinate regulation of the Suf and Isc Fe-S cluster biogenesis pathways by IscR is essential for viability of Escherichia coli. J Bacteriol 196:4315-23
Park, Dan M; Akhtar, Md Sohail; Ansari, Aseem Z et al. (2013) The bacterial response regulator ArcA uses a diverse binding site architecture to regulate carbon oxidation globally. PLoS Genet 9:e1003839
Myers, Kevin S; Yan, Huihuang; Ong, Irene M et al. (2013) Genome-scale analysis of escherichia coli FNR reveals complex features of transcription factor binding. PLoS Genet 9:e1003565

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