Binding of the transcription activator NRI (NTRC) to a supercoiled DNA segment imitates association with the natural enhancer: an electron microscopic investigation.
Electron microscopic visualization indicates that the transcription activator NRI (NTRC) binds with exceptional selectivity and efficiency to a sequence-induced superhelical (spiral) segment inserted upstream of the glnA promoter, accounting for its observed ability to substitute for the natural glnA enhancer. The cooperative binding of NRI to the spiral insert leads to protein oligomerization which, at higher concentration, promotes selective coating of the entire superhelical segment with protein. Localization of NRI at apical loops is observed with negatively supercoiled plasmid DNA. With a linear plasmid, bending of DNA is observed. We confirm that NRI is a DNA-bending protein, consistent with its high affinity for spiral DNA. These results prove that spiral DNA without any homology to the NRI-binding sequence site can substitute for the glnA enhancer by promoting cooperative activator binding to DNA and facilitating protein oligomerization. Similar mechanisms might apply to other prokaryotic and eukaryotic activator proteins that share the ability to bend DNA and act efficiently as multimers.
Révet
B
B
Laboratoire de Microscopie Cellulaire et Moléculaire (Centre National de la Recherche Scientifique, Unité de Recherche Associée 147), Institut Gustave-Roussy, Villejuif, France.
Brahms
S
S
Brahms
G
G
D001426:Bacterial Proteins; D001483:Base Sequence; D001665:Binding Sites; D004278:DNA, Superhelical / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure*; D004742:Enhancer Elements, Genetic; D066298:In Vitro Techniques; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D051922:PII Nitrogen Regulatory Proteins; D010957:Plasmids / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D011487:Protein Conformation; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure*; D014157:Transcription Factors; D015533:Transcriptional Activation
D001426:Bacterial Proteins; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D051922:PII Nitrogen Regulatory Proteins; D015534:Trans-Activators; D014157:Transcription Factors