A multiprotein binding interface in an intrinsically disordered region of the tumor suppressor protein interferon regulatory factor-1

TY - JOUR

T1 - A multiprotein binding interface in an intrinsically disordered region of the tumor suppressor protein interferon regulatory factor-1

AU - Narayan, Vikram

AU - Halada, Petr

AU - Hernychová, Lenka

AU - Chong, Yuh Ping

AU - Žáková, Jitka

AU - Hupp, Ted R

AU - Vojtesek, Borivoj

AU - Ball, Kathryn L

PY - 2011/4/22

Y1 - 2011/4/22

N2 - The interferon-regulated transcription factor and tumor suppressor protein IRF-1 is predicted to be largely disordered outside of the DNA-binding domain. One of the advantages of intrinsically disordered protein domains is thought to be their ability to take part in multiple, specific but low affinity protein interactions; however, relatively few IRF-1-interacting proteins have been described. The recent identification of a functional binding interface for the E3-ubiquitin ligase CHIP within the major disordered domain of IRF-1 led us to ask whether this region might be employed more widely by regulators of IRF-1 function. Here we describe the use of peptide aptamer-based affinity chromatography coupled with mass spectrometry to define a multiprotein binding interface on IRF-1 (Mf2 domain; amino acids 106-140) and to identify Mf2-binding proteins from A375 cells. Based on their function as known transcriptional regulators, a selection of the Mf2 domain-binding proteins (NPM1, TRIM28, and YB-1) have been validated using in vitro and cell-based assays. Interestingly, although NPM1, TRIM28, and YB-1 all bind to the Mf2 domain, they have differing amino acid specificities, demonstrating the degree of combinatorial diversity and specificity available through linear interaction motifs.

AB - The interferon-regulated transcription factor and tumor suppressor protein IRF-1 is predicted to be largely disordered outside of the DNA-binding domain. One of the advantages of intrinsically disordered protein domains is thought to be their ability to take part in multiple, specific but low affinity protein interactions; however, relatively few IRF-1-interacting proteins have been described. The recent identification of a functional binding interface for the E3-ubiquitin ligase CHIP within the major disordered domain of IRF-1 led us to ask whether this region might be employed more widely by regulators of IRF-1 function. Here we describe the use of peptide aptamer-based affinity chromatography coupled with mass spectrometry to define a multiprotein binding interface on IRF-1 (Mf2 domain; amino acids 106-140) and to identify Mf2-binding proteins from A375 cells. Based on their function as known transcriptional regulators, a selection of the Mf2 domain-binding proteins (NPM1, TRIM28, and YB-1) have been validated using in vitro and cell-based assays. Interestingly, although NPM1, TRIM28, and YB-1 all bind to the Mf2 domain, they have differing amino acid specificities, demonstrating the degree of combinatorial diversity and specificity available through linear interaction motifs.

U2 - 10.1074/jbc.M110.204602

DO - 10.1074/jbc.M110.204602

M3 - Article

C2 - 21245151

SN - 0021-9258

VL - 286

SP - 14291

EP - 14303

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 16

ER -