Determinants that control the specific interactions between TAB1 and p38alpha.
Previous studies have revealed that transforming growth factor-beta-activated protein kinase 1 (TAB1) interacts with p38alpha and induces p38alpha autophosphorylation. Here, we examine the sequence requirements in TAB1 and p38alpha that drive their interaction. Deletion and point mutations in TAB1 reveal that a proline residue in the C terminus of TAB1 (Pro412) is necessary for its interaction with p38alpha. Furthermore, a cryptic D-domain-like docking site was identified adjacent to the N terminus of Pro412, putting Pro412 in the phi(B)+3 position of the docking site. Through mutational analysis, we found that the previously identified hydrophobic docking groove in p38alpha is involved in this interaction, whereas the CD domain and ED domain are not. Furthermore, chimeric analysis with p38beta (which does not bind to TAB1) revealed a previously unidentified locus of p38alpha comprising Thr218 and Ile275 that is essential for specific binding of p38alpha to TAB1. Converting either of these residues to the corresponding amino acid of p38beta abolishes p38alpha interaction with TAB1. These p38alpha mutants still can be fully activated by p38alpha upstream activating kinase mitogen-activated protein kinase kinase 6, but their basal activity and activation in response to some extracellular stimuli are reduced. Adjacent to Thr218 and Ile275 is a site where large conformational changes occur in the presence of docking-site peptides derived from p38alpha substrates and activators. This suggests that TAB1-induced autophosphorylation of p38alpha results from conformational changes that are similar but unique to those seen in p38alpha interactions with its substrates and activating kinases.
Zhou
Huamin
H
The Key Laboratory of the Ministry of Education for Cell Biology and Tumor Cell Engineering, School of Life Sciences, Xiamen University, Fujian, China.
Zheng
Min
M
Chen
Jianming
J
Xie
Changchuan
C
Kolatkar
Anand R
AR
Zarubin
Tyler
T
Ye
Zhiyun
Z
Akella
Radha
R
Lin
Shengcai
S
Goldsmith
Elizabeth J
EJ
Han
Jiahuai
J
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000409:Alanine / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D015153:Blotting, Western; D018929:Cell Culture Techniques; D002460:Cell Line; D002942:Circular Dichroism; D004252:DNA Mutational Analysis; D004789:Enzyme Activation; D004926:Escherichia coli / Q000235:genetics; D017353:Gene Deletion; D017930:Genes, Reporter; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D013058:Mass Spectrometry; D048308:Mitogen-Activated Protein Kinase 14 / Q000032:analysis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017354:Point Mutation; D011233:Precipitin Tests; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000032:analysis; D017386:Sequence Homology, Amino Acid; D013057:Spectrum Analysis; D013912:Threonine / Q000737:chemistry; D014162:Transfection; D001616:beta-Galactosidase / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D011993:Recombinant Fusion Proteins; C486300:TAB1 protein, human; D013912:Threonine; D008156:Luciferases; D048308:Mitogen-Activated Protein Kinase 14; D001616:beta-Galactosidase; D000409:Alanine