Phosphorylation of microtubule-associated proteins from the ovaries of hemipteran insects by MPF and MAP kinase: possible roles in the regulation of microtubules during oogenesis

摘要

Nutritive tubes that link the developing oocytes to the nurse cells in ovarioles of hemipteran insects contain extensive arrays of microtubules. These are established, then later depolymerised, by developmentally regulated processes. Breakdown of the microtubules has previously been shown to correspond with the activation of M-phase promoting factor (MPF) and mitogen-activated protein kinase (MAP kinase), late in oogenesis, as the oocytes proceed to arrest at the first meiotic metaphase. The mechanisms that lead to the breakdown of nutritive tube microtubules are unknown. The possibility was investigated that the insect ovarian microtubules are regulated by MPF- or MAP kinase-dependent phosphorylation, focusing upon the prominent high molecular weight microtubule-associated protein (HMW MAP) enriched in this system, which is a potential target for protein kinase activity in vivo. The prominent HMW MAPs were purified from the ovaries of two species of hemipterans, Oncopeltus fasciatus and Notonecta glauca, and were shown to be substrates in vitro for the activities of MPF and MAP kinase. However, although the catalytic component of MPF (p34cdc2) is present within microtubule-rich portions of hemipteran ovarioles, it was found that neither this protein nor its regulatory partner (cyclin B) co-purify with microtubules during taxol-mediated microtubule isolation.