Involvement of mitogen-activated protein kinase in 2-hydroxyestradiol-17β-induced oocyte maturation in the catfish Heteropneustes fossilis and a note on possible interaction with protein phosphatases

Abstract

Mitogen-activated protein kinase (MAPK) was demonstrated in the postvitellogenic follicles (theca-granulosa and oocyte) of catfish by Western blotting using a polyclonal anti-rabbit serum, which recognized both ERK1 and ERK2. Two distinct protein bands resolved in the 46–48 kDa range of 12% SDS–PAGE were immunoblotted. Incubation of the follicles with 5 μM 2-OHE 2 elicited GVBD significantly in a duration-dependent manner with a concomitant increase in the expression of MAPK (ERK1 and ERK2). Densitometric analysis of the immunoblots showed significant variations in the intensity of staining. The ERK1 expression increased significantly from 6 h onwards but the changes were less pronounced. On the other hand, ERK2 registered a sharp significant increase after 3 h, which paralleled the GVBD response. The MEK inhibitor PD098059 alone did not induce GVBD. Co-incubation of the follicles with 2-OHE 2 and PD098059 significantly inhibited the steroid-induced GVBD at all concentrations. Immunoblot analysis showed that PD098059 inhibited MAPK activity significantly compared to the 2-OHE 2 group. The addition of okadaic acid (OA) in the incubation medium containing both 2-OHE 2 and PD098059 reversed the inhibitory effect of the latter and GVBD was elevated significantly over that of the 2-OHE 2 group but significantly lower than that of the 2-OHE 2 + OA group. The results suggest an involvement of MAPK in meiotic maturation but the site(s) of action: oocyte, follicular envelope or both needs further investigation.