Abstract

There is growing appreciation that small, non-coding RNAs can participate in gene regulation. Can small RNAs inhibit DNA-binding proteins? We have developed an artificial example by performing in vitro genetic selection experiments identifying a small RNA aptamer that competitively inhibits human transcription factor NF-kappaB binding to DNA in vitro. Optimization by yeast in vivo genetic selections resulted in an RNA that inhibits NF-kappaB in living yeast cells. We have solved the X-ray co-crystal structure of this unusual RNA/NF-I Four specific aims are proposed:
Aim 1. Optimize and extend anti-NF-kappaB RNA aptamer function by genetic selections in bacteria and yeast.
Aim 2. Analyze the structural basis for NF-kappaB binding by anti-NF-kappaB RNA aptamers.
Aim 3. Analyze NF-kappaB inhibition by anti-NF-kappaB RNA aptamers in cultured mammalian cells.
Aim 4. Identify small molecules that inhibit DNA binding by NF-kappaB.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM068128-04
Application #
7169628
Study Section
Special Emphasis Panel (ZRG1-CDF-1 (90))
Program Officer
Tompkins, Laurie
Project Start
2004-02-01
Project End
2009-01-31
Budget Start
2007-02-01
Budget End
2009-01-31
Support Year
4
Fiscal Year
2007
Total Cost
$279,714
Indirect Cost

  Institution

Name
Mayo Clinic, Rochester
Department
Type
DUNS #
006471700
City
Rochester
State
MN
Country
United States
Zip Code
55905

  Publications

Mondragon, Estefania; Maher 3rd, L James (2012) An ace in the hole... Structure 20:1285-6
Nastasijevic, Branislav; Wright, Brent R; Smestad, John et al. (2012) Remyelination induced by a DNA aptamer in a mouse model of multiple sclerosis. PLoS One 7:e39595
Bida, John Paul; Maher 3rd, L James (2012) Improved prediction of RNA tertiary structure with insights into native state dynamics. RNA 18:385-93
Wurster, Susan E; Maher 3rd, L James (2010) Selections that optimize RNA display in the yeast three-hybrid system. RNA 16:253-8
Wurster, Susan E; Bida, John Paul; Her, Yeng F et al. (2009) Characterization of anti-NF-kappaB RNA aptamer-binding specificity in vitro and in the yeast three-hybrid system. Nucleic Acids Res 37:6214-24
Reiter, Nicholas J; Maher 3rd, L James; Butcher, Samuel E (2008) DNA mimicry by a high-affinity anti-NF-kappaB RNA aptamer. Nucleic Acids Res 36:1227-36
Wurster, Susan E; Maher 3rd, L James (2008) Selection and characterization of anti-NF-kappaB p65 RNA aptamers. RNA 14:1037-47
Riley, Kasandra J-L; James Maher 3rd, L (2007) Analysis of p53-RNA interactions in cultured human cells. Biochem Biophys Res Commun 363:381-7
Riley, Kasandra J-L; Maher 3rd, L James (2007) p53 RNA interactions: new clues in an old mystery. RNA 13:1825-33
Riley, Kasandra J-L; Ramirez-Alvarado, Marina; Maher 3rd, L James (2007) RNA-p53 interactions in vitro. Biochemistry 46:2480-7

Showing the most recent 10 out of 11 publications