Structural basis for the selective inhibition of JNK1 by the scaffolding protein JIP1 and SP600125

Heo, Yong Seok; Kim, Su Kyoung; Seo, Chang Il; Kim, Young Kwan; Sung, B.-J.; Lee, Hye Shin; Lee, Jae Il; Park, Sam Yong; Kim, Jin Hwan; Hwang, Kwang Yeon; Hyun, Young Lan; Jeon, Young Ho; Ro, Seonggu; Cho, Joong Myung; Lee, Tae Gyu; Yang, Chu Hak

doi:10.1038/sj.emboj.7600212

Cited by 253 publications

(340 citation statements)

References 47 publications

Supporting

Mentioning

308

Contrasting

Unclassified

Order By: Relevance

“…This conclusion that Leu-131 is important in the KIM binding interface of JNK is supported further by the data published on JNK1 in complex with TI-JIP just prior to the submission of our study (23). Consistent with the models presented in Fig.…”

Section: Discussionsupporting

confidence: 79%

“…A, space-filling structure with JNK1 residues Leu-131 and Tyr-320 highlighted in purple, which were implicated in the interaction between JNK1 and the TI-JIP inhibitor. Residues that were implicated in the interaction between JNK1 and TI-JIP by co-crystallization analysis (23) are colored green. B, the hydrophobic pocket thought to interact with the L-X-L motif of TI-JIP is circled and consists of JNK1 residues Ala-113, Leu-115, Val-118, Met-121, Leu-123, Leu-131, Val-159, and Cys-163 (23).…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Reverse Two-hybrid Screening Identifies Residues of JNK Required for Interaction with the Kinase Interaction Motif of JNK-interacting Protein-1

Barr

Hopkins

Watt

et al. 2004

Journal of Biological Chemistry

View full text Add to dashboard Cite

The development of specific inhibitors for the c-Jun N-terminal kinase (JNK) family of mitogen-activated protein kinases (MAPKs) has been a recent research focus because of the association of JNK with cell death in conditions such as stroke and neurodegeneration. We have demonstrated previously the presence of critical inhibitory residues within an 11-mer peptide (TI-JIP) based on the sequence of JNK-interacting protein-1 (JIP-1). However, the corresponding region of JNK bound by this JIP-1-based peptide was unknown. To identify this region, we used a novel reverse two-hybrid approach with TI-JIP as bait. We screened a library of JNK1 mutants that had been generated by random PCR mutagenesis and found three mutants of JNK1 that failed to interact with TI-JIP. The mutations in JNK1 were L131R, R309W, and Y320H. Of these mutated residues, Leu-131 and Tyr-320 were located on a common face of the JNK protein close to other residues implicated previously in the interactions of MAPKs with substrates, phosphatases, and scaffolds. To test whether these JNK1 mutants were thus affected in their regulation, we evaluated their activation in mammalian cells in response to hyperosmolarity or cotransfection with a constitutively active upstream kinase or their direct phosphorylation by either MAPK kinase (MKK)4 or MKK7. In each situation, all three JNK mutants were not activated or phosphorylated to the same level as wildtype JNK. Therefore, the results of our unbiased reverse two-hybrid screening approach have identified residues of JNK responsible for binding JIP-1-based peptides as well as MKK4 or MKK7.The c-Jun N-terminal kinase (JNK) 1 subfamily of mitogenactivated protein kinases (MAPKs) is activated after the exposure of cells to various stimuli including growth factors, cytokines, and cellular stresses (1-5). This activation of JNK requires the phosphorylation of Thr and Tyr residues in its activation loop by the upstream kinases MKK4 and MKK7 (6). Activated JNK then phosphorylates nuclear substrates including c-Jun, ATF-2, and Elk-1 and non-nuclear substrates such as Bcl-2 family members. This allows JNK to contribute to diverse biological processes including cell proliferation, differentiation, survival, and death (for review, see Ref. 7).Selective JNK or JNK pathway inhibitors have been developed after the implication of JNK signaling in various disease states. Presently, there are three classes of JNK pathway inhibitors. The first of these is Cephalon Incorporated library compound 1347 (CEP-1347), which inhibits the mixed lineage kinases that function upstream in the JNK pathway (8). This compound inhibits JNK activation both in vitro and in vivo (9 -12) and is currently in phase II clinical trials for the treatment of Parkinson's disease. The second inhibitor is Signal Pharmaceuticals library compound 600125 (SP600125), which is a reversible ATP-competitive JNK inhibitor (13). However, the selectivity of this compound for JNK is questionable after recent specificity tests where SP600125 inhibited 13 other protein ...

show abstract

Section: Discussionsupporting

confidence: 79%

Section: Discussionmentioning

confidence: 99%

Reverse Two-hybrid Screening Identifies Residues of JNK Required for Interaction with the Kinase Interaction Motif of JNK-interacting Protein-1

Barr

Hopkins

Watt

et al. 2004

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…The DRS of ERK2 recognizes a D-site either with the sequence of (R/K) 2-3 -X 4-6 -Φ A -X-Φ B (in this sequence X represents any amino acid and Φ = Leu, Ile, or Val) or a related sequence found in MAPKAP kinases (11). The structures of several D-sites bound to the DRSs of the three major MAPK classes (p38 MAPK (13), JNK (14), and ERK2 (15,16)) have been reported. These structures reveal a common, but not uniform, mode of binding where the characteristic basic residues engage acidic residues on loop-16, while the Φ residues bind to hydrophobic sites (indicated by Ø) within the groove (See Figures 2A and 2B which depicts two D-sites bound to ERK2 (15,16)).…”

mentioning

confidence: 99%

Expanding the Repertoire of an ERK2 Recruitment Site: Cysteine Footprinting Identifies the D-Recruitment Site as a Mediator of Ets-1 Binding

et al. 2007

View full text Add to dashboard Cite

Many substrates of ERK2 contain a D-site, a sequence recognized by ERK2 that is used to promote catalysis. Despite lacking a canonical D-site, the substrate Ets-1 is displaced from ERK2 by peptides containing one. This suggests that Ets-1 may contain a novel or cryptic D-site. To investigate this possibility a protein footprinting strategy was developed to elucidate ERK2-ligand interactions. Using this approach, single cysteine reporters were placed in the D-recruitment site (DRS) of ERK2 and the resulting ERK2 proteins subjected to alkylation by iodoacetamide. The ability of residues 1-138 of Ets-1 to protect the cysteines from alkylation was determined. The pattern of protection observed is consistent with Ets-1 occupying a hydrophobic binding site within the DRS of ERK2. Significantly, a peptide derived from the D-site of Elk-1, which is known to bind the DRS, exhibits a similar pattern of cysteine protection. This analysis expands the repertoire of the DRS on ERK2 and suggests that other targeting sequences remain to be identified. Furthermore, cysteinefootprinting is presented as a useful way to interrogate protein-ligand interactions at the resolution of a single amino acid.The mitogen-activated protein kinases (MAPKs) 1 are ubiquitous elements of eukaryotic signaling pathways that permeate nearly all aspects of signaling in multicellular organisms. In humans the loss of their regulation is associated with many diseases that encompass an expanding list of cancers as well as neurological and inflammatory diseases (1). Three main subgroups have been identified in humans, termed the ERKs (2), the JNKs (3,4), and the p38 MAPKs (5,6). Several newer members have recently been reviewed (7). Extracellular regulated protein kinase 2 (ERK2), the prototypical member of the MAP kinase family, catalyzes the † This research was supported in part by the Welch Foundation (F-1390) and the National Institutes of Health (GM59802). Mass spectra were acquired by Dr. Herng-Hsiang Lo in the CRED Analytical Instrumentation Facility Core supported by the NIEHS center grant ES07784. Molecular graphics images were produced using the UCSF Chimera package from the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco (supported by NIH P41 RR-01081). NIH Public Access Author ManuscriptBiochemistry. Author manuscript; available in PMC 2010 July 6. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript transfer of the γ-phosphate of adenosine triphosphate to serine or threonine residues that generally lie in flexible regions of proteins. It is a remarkable enzyme whose ability to exhibit high specificity toward a discrete subset of structurally diverse proteins is poorly understood.Protein kinases generally recognize substrates through a consensus sequence that contains the phosphorylation site (8). This sequence is recognized, in part, by a region called the activation segment that encompasses residues 165 DFG to APE 195 of the catalytic domain of protein ki...

show abstract

“…Structural and mutagenesis studies indicate that the basic submotif of the D-site contacts two closely spaced acidic patches on the MAPK surface-the 'CD/sevenmaker' region and the 'ED region' [24,[54][55][56][57][58]. The hydrophobic submotif of the D-site binds to a 'hydrophobic docking groove' located close the CD/ED regions [54,56,57,59].…”

Section: Cognate Docking Sites On Mapksmentioning

confidence: 99%

“…The hydrophobic submotif of the D-site binds to a 'hydrophobic docking groove' located close the CD/ED regions [54,56,57,59].…”

Section: Cognate Docking Sites On Mapksmentioning

confidence: 99%

Analysis of mitogen-activated protein kinase activation and interactions with regulators and substrates

Bardwell

Shah

2006

Methods

View full text Add to dashboard Cite

Mitogen-activated protein kinase (MAPK) cascades are ubiquitous signal transduction modules in eukaryotes that are of great interest and importance. Here, we summarize some useful methods for the analysis of MAPK signaling, including methods to (1) detect MAPK activation in cells, with an emphasis on using phosphorylation-state-specific antibodies raised against mammalian phosphopeptide sequences to detect the activation of MAPKs in other species; (2) estimate the cellular concentrations of MAPKs and other proteins of interest; (3) detect and quantify the stable physical association of MAPKs with their substrates and regulators, and estimate the relevant dissociation constants; (4) delineate the MAPK-binding regions or domains of MAPK-interacting proteins, with particular emphasis on the identification and verification of MAPK-docking sites. These procedures are broadly applicable to many organisms, including both yeast and mammalian cells.

show abstract

Structural basis for the selective inhibition of JNK1 by the scaffolding protein JIP1 and SP600125

Cited by 253 publications

References 47 publications

Reverse Two-hybrid Screening Identifies Residues of JNK Required for Interaction with the Kinase Interaction Motif of JNK-interacting Protein-1

Reverse Two-hybrid Screening Identifies Residues of JNK Required for Interaction with the Kinase Interaction Motif of JNK-interacting Protein-1

Expanding the Repertoire of an ERK2 Recruitment Site: Cysteine Footprinting Identifies the D-Recruitment Site as a Mediator of Ets-1 Binding

Analysis of mitogen-activated protein kinase activation and interactions with regulators and substrates

Contact Info

Product

Resources

About