[Heat shock activated Rac-MEKK-JNK pathway and hsp90 beta gene expression].
OBJECTIVE
To study the effect of Rac-MEKK-JNK (Rac-mitogen activated protein kinase kinase kinase-C-jun N-terminal protein kinase) signal pathway on heat shock-induced hsp90 beta gene expression and the impact of Hsp90 on the regulation of the pathway.


METHODS
DN-Rac, DN-MEKK or DN-JNK were cotransfected with hsp90 beta CAT reporter plasmid beta 3.1 into Jurkat or LETPa-2 cells individually, the CAT mRNA expression was then determined quantitatively by competitive RT-PCR based system. Western blot was carried out to detect the expression level and phosphorylation of c-Jun in Jurkat and LETPa-2 cells that were transfected with DN-Rac, DN-MEKK or DN-JNK. By in vitro kinase activity assay and Western blot, the effect of geldnamycin (GA) on heat induced JNK activity were evaluated.


RESULTS
In Jurkat cell transfected with DN-Rac, DN-MEKK or DN-JNK, heat shock induced relative CAT mRNA expression level was decreased to (72.8 +/- 5)%, (60 +/- 13.2)% and (47.7 +/- 12.1)% of the control respectively; while in LETPa-2 cell hsp90 beta 3.1 reporter gene expression was accordingly suppressed to (16.17 +/- 5.1)%, (50.2 +/- 8.7)% and (47.5 +/- 10)% of control. C-Jun expression and phosphorylation were inhibited by the transfection of either one of DN-Rac, DN-MEKK or DN-JNK. With GA treatment, heat shock induced JNK activity was repressed, while the expression level of JNK or c-Jun was not obviously changed.


CONCLUSIONS
Rac-MEKK-JNK pathway promotes heat shock induced hsp90 beta gene expression and hsp90 may participate in the regulation of heat shock activated Rac-MEKK-JNK signal pathway in both Jurkat and LETPa-2 cells.
Li
Xiao-yan
XY
National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
Lu
Cheng
C
Wu
Ning-hua
NH
Shen
Yu-fei
YF
D016227:Benzoquinones; D045744:Cell Line, Tumor; D017930:Genes, Reporter; D018841:HSP90 Heat-Shock Proteins / Q000096:biosynthesis* / Q000235:genetics; D006358:Hot Temperature; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases; D047029:Lactams, Macrocyclic; D015458:Leukemia, T-Cell / Q000473:pathology; D020929:Mitogen-Activated Protein Kinase Kinases / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology*; D011493:Protein Kinase C / Q000502:physiology; D011809:Quinones / Q000494:pharmacology; D015398:Signal Transduction; D014162:Transfection
D016227:Benzoquinones; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D011809:Quinones; D011493:Protein Kinase C; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; C001277:geldanamycin