title: Functional interactions of Raf and MEK with Jun-N-terminal kinase (JNK) result in a positive feedback loop on the oncogenic Ras signaling pathway.
abstract: In previous studies we have found that oncogenic (Val 12)-ras-p21 induces Xenopus laevis oocyte maturation that is selectively blocked by two ras-p21 peptides, 35-47, also called PNC-7, that blocks its interaction with raf, and 96-110, also called PNC-2, that blocks its interaction with jun-N-terminal kinase (JNK). Each peptide blocks activation of both JNK and MAP kinase (MAPK or ERK) suggesting interaction between the raf-MEK-ERK and JNK-jun pathways. We further found that dominant negative raf blocks JNK induction of oocyte maturation, again suggesting cross-talk between pathways. In this study, we have undertaken to determine where these points of cross-talk occur. First, we have immunoprecipitated injected Val 12-Ha-ras-p21 from oocytes and found that a complex forms between ras-p21 raf, MEK, MAPK, and JNK. Co-injection of either peptide, but not a control peptide, causes diminished binding of ras-p21, raf, and JNK. Thus, one site of interaction is cooperative binding of Val 12-ras-p21 to raf and JNK. Second, we have injected JNK, c-raf, and MEK into oocytes alone and in the presence of raf and MEK inhibitors and found that JNK activation is independent of the raf-MEK-MAPK pathway but that activated JNK activates raf, allowing for activation of ERK. Furthermore, we have found that constitutively activated MEK activates JNK. We have corroborated these findings in studies with isolated protein components from a human astrocyte (U-251) cell line; that is, JNK phosphorylates raf but not the reverse; MEK phosphorylates JNK but not the reverse. We further have found that JNK does not phosphorylate MAPK and that MAPK does not phosphorylate JNK. The stress-inducing agent, anisomycin, causes activation of JNK, raf, MEK, and ERK in this cell line; activation of JNK is not inhibitable by the MEK inhibitor, U0126, while activation of raf, MEK, and ERK are blocked by this agent. These results suggest that activated JNK can, in turn, activate not only jun but also raf that, in turn, activates MEK that can then cross-activate JNK in a positive feedback loop.
authors.affiliation: QRNA Corporation, West 167th Street, New York, New York 10032, USA.
authors.forename: Victor
authors.initials: V
authors.lastname: Adler
authors.forename: Yongxia
authors.initials: Y
authors.lastname: Qu
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Smith
authors.forename: Lara
authors.initials: L
authors.lastname: Izotova
authors.forename: Sidney
authors.initials: S
authors.lastname: Pestka
authors.forename: Hsiang-Fu
authors.initials: HF
authors.lastname: Kung
authors.forename: Marie
authors.initials: M
authors.lastname: Lin
authors.forename: Fred K
authors.initials: FK
authors.lastname: Friedman
authors.forename: Lyndon
authors.initials: L
authors.lastname: Chie
authors.forename: Denise
authors.initials: D
authors.lastname: Chung
authors.forename: Mohamed
authors.initials: M
authors.lastname: Boutjdir
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Pincus
mesh_terms: D000818:Animals; D002070:Butadienes / Q000494:pharmacology; D045744:Cell Line, Tumor; D025461:Feedback, Physiological; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D048369:MAP Kinase Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D046912:Multiprotein Complexes; D009570:Nitriles / Q000494:pharmacology; D009865:Oocytes; D010766:Phosphorylation; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism*; D020239:Receptor Cross-Talk; D015398:Signal Transduction; D014982:Xenopus laevis
chemical_list: D002070:Butadienes; D046912:Multiprotein Complexes; D009570:Nitriles; C113580:U 0126; D019908:Proto-Oncogene Proteins c-raf; D048031:JNK Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)