טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentDaniliuc Sharon
SubjectThe Identification and Characterization of a New c-Jun
N-terminal Kinase (JNK) Binding Protein
DepartmentDepartment of Medicine
Supervisor Professor Ami Aronheim
Full Thesis textFull thesis text - English Version


Abstract

Mitogen- activated protein kinases (MAPKs) regulate a variety of cellular processes in response to extracellular signals. MAPKs are activated through a protein kinase cascade. c-Jun N-terminal kinase (JNK), a stress activated MAPK protein is one of the three MAPK members in mammals. The activation of the JNK cascade results in changes in the pattern of gene expression and diverse biological responses. Efficient signaling is mediated by scaffold proteins that simultaneously associate with various components of the MAPK signaling pathway. In this study we isolated a new scaffold protein for the JNK module using the yeast RRS system, and named it JNK binding protein; JBP. JBP is part of the WDR62 gene (accession 043379-1). We hypothesize that JBP serves as a new scaffold for JNK. The association between JNK and JBP was confirmed in mammalian cells as JBP was able to co-precipitate the over expression proteins of JNK1 and JNK2 as well as MKK7. The C-terminal of JBP is crucial for its association with JNK. JBP over expression, with no additional stimuli, potentiates JNK2 kinase activity by 15 fold as well as JNK1 kinase activity by 9 fold in a saturable dependent manner. This potentiation of activity is specific to the JNK family. This activation does not result in the activation of AP-1 dependent transcription. Using immuno-fluorescence JBP staining was found to be localized to stress granules (SG). SG are granules that are formed following cell stress to protect mRNA from degradation. This unique JNK localization requires the interaction between JBP and may suggest a role of JNK in mRNA determination process. In addition, endogenous WDR62 is found at the center of the duplicated centrosome. JBP appearance at the centrosome lasts from the beginning of mitosis at prophase till telophase but is absent from interphase stage. We also found that JBP is a bona fide JNK substrate in vitro. JBP phosphorylation is mapped to the six SP potential JNK phosphorylation sites located at the C-terminus. 

Collectively, we propose that JBP/WDR62 is a new scaffold protein for the JNK module, located to stress granules in the cell and directing a non-classical JNK activation to SG and processing bodies. JBP is able to activate JNK in a dose dependent manner. This new sub-cellular location of JNK activity in the cell during stress, may contribute to a new level of regulation of stalled mRNA following cell stress.