NonO enhances the association of many DNA-binding proteins to their targets.
NonO is an unusual nucleic acid binding protein not only in that it binds both DNA and RNA but that it does so via functionally separable domains. Here we document that NonO enhances the binding of some (E47, OTF-1 and OTF-2) but not all (PEA3) conventional sequence-specific transcription factors to their recognition sites in artificial substrates as well as in an immunoglobulin VHpromoter. We also show that NonO induces the binding of the Ku complex to DNA ends. Ku has no known DNA sequence specificity. These enhancement of binding effects are NonO concentration dependent. Using the E box activity of E47 as a model, kinetic studies demonstrate that the association rate of the protein-DNA complex increases in the presence of NonO while the dissociation rate remains the same, thereby increasing the sum total of the interaction. Oligo competition experiments indicate that NonO does not contact the target DNA in order to enhance the binding activity of DNA binding proteins. Rather, methylation interference analysis reveals that the induced E47 binding-activity has the same DNA-binding sequence specificity as the normal binding. This result suggests that one of the effects of NonO is to induce a true protein-DNA interaction. In this way, it might be possible for NonO to play a crucial role in gene regulation.
Yang
Y S
YS
Molecular Immunology Center, Department of Microbiology, The University of Texas Southwestern Medical Center, 6000 Harry Hines Boulevard, Dallas, TX 75235-9140, USA.
Yang
M C
MC
Tucker
P W
PW
Capra
J D
JD
D034961:Antigens, Nuclear; D001665:Binding Sites; D019938:Cyclin D1; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004265:DNA Helicases; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000378:metabolism*; D005786:Gene Expression Regulation; D005803:Genes, Immunoglobulin; D005982:Glutathione Transferase; D051863:Host Cell Factor C1; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007135:Immunoglobulin Variable Region / Q000235:genetics; D007700:Kinetics; D000072200:Ku Autoantigen; D009687:Nuclear Proteins / Q000378:metabolism; D050811:Octamer Transcription Factor-1; D015345:Oligonucleotide Probes; D010843:Pichia; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000096:biosynthesis / Q000378:metabolism*; D014158:Transcription, Genetic
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D051863:Host Cell Factor C1; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D009687:Nuclear Proteins; D050811:Octamer Transcription Factor-1; D015345:Oligonucleotide Probes; D011518:Proto-Oncogene Proteins; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C081647:high affinity DNA-binding factor, S cerevisiae; D019938:Cyclin D1; D004247:DNA; D005982:Glutathione Transferase; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen