title: Replacement of the C-terminal tetrapeptide (314 PAPV 317 to 314 SSSM 317) in interferon regulatory factor-2 alters its N-terminal DNA-binding activity.
abstract: Interferon regulatory factor-2 (IRF-2) is an important transcription factor involved in cell growth regulation, immune response and cancer. IRF-2 can function as a transcriptional repressor and activator depending on its DNA-binding activity and protein-protein interactions. We compared the amino acid sequences of IRF-2 and found a C-terminal tetrapeptide (314PAPV317) of mouse IRF-2 to be different (314SSSM317) from human IRF-2. Recombinant GST-IRF-2 with 314PAPV317 (wild type) and 314SSSM317 (mutant) expressed in Escherichia coli were assessed for DNA-binding activity with 32P-(GAAAGT) 4 by electrophoretic mobility shift assay (EMSA). Wild type- and mutant GST-IRF-2 showed similar expression patterns and immunoreactivities but different DNA-binding activities. Mutant (mt) IRF-2 formed higher-molecular-mass, more and stronger DNA-protein complexes in comparison to wild type (wt) IRF-2. Anti-IRF-2 antibody stabilized the DNA-protein complexes formed by both wt IRF-2 and mt IRF-2, resolving the differences. This suggests that PAPV and SSSM sequences at 314-317 in the C-terminal region of mouse and human IRF-2 contribute to conformation of IRF-2 and influence DNA-binding activity of the N-terminal region, indicating intramolecular interactions. Thus, evolution of IRF-2 from murine to human genome has resulted in subtle differences in C-terminal amino acid motifs, which may contribute to qualitative changes in IRF-2-dependent DNA-binding activity and gene expression.
authors.affiliation: Molecular Biology Laboratory, School of Life Sciences, Jawaharlal Nehru University, New Delhi 110 067, India.
authors.forename: Krishna
authors.initials: K
authors.lastname: Prakash
authors.forename: Pramod C
authors.initials: PC
authors.lastname: Rath
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001667:Binding, Competitive; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D050836:Interferon Regulatory Factor-2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009842:Oligopeptides / Q000737:chemistry*; D011119:Polynucleotides / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D004268:DNA-Binding Proteins; C494229:IRF2 protein, human; D050836:Interferon Regulatory Factor-2; C494230:Irf2 protein, mouse; D009842:Oligopeptides; D011119:Polynucleotides; D011993:Recombinant Fusion Proteins