Phospholipid vesicle binding and aggregation by four novel fish annexins are differently regulated by Ca2+.

Four members of the annexin family, herein referred to as max (for medaka annexin) 1-4, have recently been identified through hybridization cloning in the killifish Oryzias latipes (D. Osterloh, J. Wittbrodt and V. Gerke, Characterization and developmentally regulated expression of four annexins in the killifish medaka. DNA and Cell Biol., in press). These annexins which are expressed in a developmentally regulated manner are present as a maternal pool in unfertilized eggs of another fish species, Misgurnus fossilis, and it has been proposed that they play a role in the Ca2+-regulated exocytosis of cortical granules occurring after fertilization. To characterize biochemical properties of the medaka proteins possibly relevant to their function in early development, we analyzed the ability of recombinantly expressed max 1-4 to interact with the principal structures of the egg cortex, phospholipid membranes and actin filaments. We show that all medaka annexins bind to acidic phospholipids in a Ca2+-regulated manner, although exhibiting different Ca2+ sensitivities. All medaka annexins, but max 1, are also capable of inducing, in a Ca2+-dependent manner, phospholipid vesicle aggregation, albeit only max 3 displays this activity at Ca2+ concentrations met in stimulated (i.e. fertilized) eggs. Max 3 is also the only medaka annexin able to interact with F-actin in the presence of Ca2+. These data identify by biochemical criteria max 3 as a close relative of the mammalian annexins I and II, thus supporting previous sequence-based comparisons. Max 3 is therefore the prime annexin candidate for being involved in cortical granule exocytosis, possibly by providing granule granule, granule plasma membrane and/or granule cytoskeleton contacts.