Molecular identification and functional characterization of a Drosophila dual-specificity phosphatase DMKP-4 which is involved in PGN-induced activation of the JNK pathway.
MAP (Mitogen-activated protein) kinases play an important role in regulating many critical cellular processes. The inactivation of MAP kinases is always accomplished by a family of dual-specificity phosphatases, termed MAPK phosphatases (MKPs). Here, we have identified a novel MKP-like protein, designated DMKP-4, from the Drosophila genome. DMKP-4 is a protein of 387 amino acids, with a dual-specificity phosphatase (DSP) catalytic domain. Recombinant protein DMKP-4 retains intrinsic phosphatase activity against chromogenic substrate pNPP. Overexpression of DMKP-4 inhibited the activation of ERK, JNK and p38 by H(2)O(2), sorbitol and heat shock in HEK293-T cells, and JNK activation in Drosophila S2 cells under PGN stimuli. "Knockdown" of DMKP-4 expression by RNAi significantly enhanced the PGN-stimulated activation of JNK, but not ERK nor p38. Further study revealed that DMKP-4 interacted specifically with JNK via its DSP domain. Mutation of Cys-126 to serine in the DSP domain of DMKP-4 not only eliminated its interaction with JNK, but also markedly reduced its phosphatase activity. Thus, DMKP-4 is a Drosophila homologue of mammalian MKPs, and may play important roles in the regulation of various developmental processes.
Sun
Lei
L
Laboratory of Signal Transduction, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, 225 South Chongqing Road, Shanghai 200025, China.
Yu
Ming-Can
MC
Kong
Ling
L
Zhuang
Zi-Heng
ZH
Hu
Jun-Hao
JH
Ge
Bao-Xue
BX
D000595:Amino Acid Sequence; D000814:Aniline Compounds / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003545:Cysteine / Q000235:genetics; D029721:Drosophila Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000166:cytology / Q000201:enzymology*; D054637:Dual-Specificity Phosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D006367:HeLa Cells; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009943:Organophosphorus Compounds / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D015398:Signal Transduction; D014162:Transfection
D000814:Aniline Compounds; D029721:Drosophila Proteins; D009943:Organophosphorus Compounds; C068798:4-aminophenylphosphate; D048031:JNK Mitogen-Activated Protein Kinases; D054637:Dual-Specificity Phosphatases; D003545:Cysteine