PARENT SESSION
Signal Transduction
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ROLE OF C-JUN N-TERMINAL KINASE AND AP-1 FACTOR IN OVOTOXICITY INDUCED BY 4-VINYLCYCLOHEXENE DIEPOXIDE IN RATS.
Hu, Xiaoming1, Sipes, I Glenn2, 3, Hoyer, Patricia1, 3, 1 2 3
ABSTRACT- Previous studies have shown that ovotoxicity induced in rats by 4-vinylcyclohexene diepoxide (VCD) is via acceleration of the normal rate of atresia. This acceleration is associated with alterations in caspase cascades, and sub-cellular distribution of Bcl-2 family members. C-Jun, a major component of the heterodimeric transcription factor AP-1, is involved in cell growth and differentiation. C-Jun can be modified by N-terminal phosphorylation (JNP) through Jun N-terminal kinase (JNK) which is associated with apoptosis. The relationship between c-Jun phosphorylation and AP-1 binding is not well understood. This study investigated the effect of VCD dosing in rats on JNK, JNP, and AP-1 activity in isolated small ovarian follicles. Female F344 rats were given a single dose of VCD (80 mg/kg, i.p., 1d; a time when ovotoxicity has not been initiated), or dosed daily for 15 days (80 mg/kg, i.p., 15d; a time when significant ovotoxicity is underway). 4 h following the final dose, livers and ovaries were collected. Ovarian small pre-antral follicles (25-100 
m) were isolated, and sub-cellular fractions were prepared for analysis. Compared with control, follicular cytosolic JNK phosphotransferase activity was decreased (~30%) by 1d of VCD dosing, but increased by 15d dosing (1.64 ± 0.38, VCD/Control, p<0.05). JNP levels in small ovarian follicles were unchanged on 1d, but increased after 15d of VCD (1.22 ± 0.1, VCD/Control, p<0.05), with no change in total c-Jun protein levels. AP-1 binding activity of follicular nuclear extracts was increased on 1d of dosing with VCD (1.43 ± 0.31, VCD/Control, p<0.05), and decreased on 15d (0.67 ± 0.08, VCD/Control, p<0.05). VCD caused no effect in liver on any measurement. Using antibodies against c-Jun or phospho-c-Jun, super-shift DNA binding demonstrated that c-Jun is contained in the AP-1-DNA binding complex. This binding is reduced with increased phospho-c-Jun. Taken together, these data provide evidence that accelerated atresia induced by VCD is associated with increased JNK activity and phosphorylation of c-Jun in rat small ovarian follicles. Because phospho-c-Jun reduces AP-1 binding, this increased phosphorylation may reduce AP-1 transcriptional activity and, in that way, enhance apoptosis (ES09246; ES06694).
KEY WORDS: small ovarian follicle, 4-vinylcyclohexene diepoxide , c-Jun N-terminal kinase, activator protein-1 transcription factor