Unknown

Dataset Information

0

Protein Cofactors Are Essential for High-Affinity DNA Binding by the Nuclear Factor ?B RelA Subunit.


ABSTRACT: Transcription activator proteins typically contain two functional domains: a DNA binding domain (DBD) that binds to DNA with sequence specificity and an activation domain (AD) whose established function is to recruit RNA polymerase. In this report, we show that purified recombinant nuclear factor ?B (NF-?B) RelA dimers bind specific ?B DNA sites with an affinity significantly lower than that of the same dimers from nuclear extracts of activated cells, suggesting that additional nuclear cofactors might facilitate DNA binding by the RelA dimers. Additionally, recombinant RelA binds DNA with relatively low affinity at a physiological salt concentration in vitro. The addition of p53 or RPS3 (ribosomal protein S3) increases RelA:DNA binding affinity 2- to >50-fold depending on the protein and ionic conditions. These cofactor proteins do not form stable ternary complexes, suggesting that they stabilize the RelA:DNA complex through dynamic interactions. Surprisingly, the RelA-DBD alone fails to bind DNA under the same solution conditions even in the presence of cofactors, suggesting an important role of the RelA-AD in DNA binding. Reduced RelA:DNA binding at a physiological ionic strength suggests that multiple cofactors might be acting simultaneously to mitigate the electrolyte effect and stabilize the RelA:DNA complex in vivo. Overall, our observations suggest that the RelA-AD and multiple cofactor proteins function cooperatively to prime the RelA-DBD and stabilize the RelA:DNA complex in cells. Our study provides a mechanism for nuclear cofactor proteins in NF-?B-dependent gene regulation.

SUBMITTER: Mulero MC 

PROVIDER: S-EPMC5993198 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

altmetric image

Publications

Protein Cofactors Are Essential for High-Affinity DNA Binding by the Nuclear Factor κB RelA Subunit.

Mulero Maria Carmen MC   Shahabi Shandy S   Ko Myung Soo MS   Schiffer Jamie M JM   Huang De-Bin DB   Wang Vivien Ya-Fan VY   Amaro Rommie E RE   Huxford Tom T   Ghosh Gourisankar G  

Biochemistry 20180510 20


Transcription activator proteins typically contain two functional domains: a DNA binding domain (DBD) that binds to DNA with sequence specificity and an activation domain (AD) whose established function is to recruit RNA polymerase. In this report, we show that purified recombinant nuclear factor κB (NF-κB) RelA dimers bind specific κB DNA sites with an affinity significantly lower than that of the same dimers from nuclear extracts of activated cells, suggesting that additional nuclear cofactors  ...[more]

Similar Datasets

| S-EPMC5704467 | biostudies-literature
| S-EPMC3152356 | biostudies-literature
| S-EPMC121441 | biostudies-literature
2015-03-31 | E-GEOD-65721 | biostudies-arrayexpress
2015-03-31 | GSE65721 | GEO
2015-03-31 | E-GEOD-65690 | biostudies-arrayexpress
2015-03-31 | E-GEOD-65707 | biostudies-arrayexpress
2015-03-31 | GSE65707 | GEO
2015-03-31 | GSE65690 | GEO
| S-EPMC4494785 | biostudies-literature