The late pollen-specific actins in angiosperms

The actin gene family of Arabidopsis has eight functional genes that are gr ouped into two ancient classes, vegetative and reproductive, and into five subclasses based on their phylogeny and mRNA expression patterns. Progress in deciphering the functional significance of this diversity is hindered by the lack of tools that can distinguish the highly conserved subclasses of actin proteins at the biochemical and cellular level. In order to address t he functional diversity of actin isovariants, we have used Arabidopsis reco mbinant actins as immunogens and produced several new anti-actin monoclonal antibodies. One of them, MAb45a, specifically recognizes two closely relat ed reproductive subclasses of actins. On immunoblots, MAb45a reacts strongl y with actins expressed in mature pollen but not with actins in other Arabi dopsis tissues. Moreover, immunocytochemical studies show that this antibod y can distinguish actin filaments in pollen tubes from those in most vegeta tive tissues. Peptide competition analyses demonstrate that asparagine at p osition 79 (Asn79) within an otherwise conserved sequence is essential for MAb45a specificity. Actins with the Asn79 epitope are also expressed in the mature pollen from diverse angiosperms and Ephedra but not from lower gymn osperms, suggesting that this epitope arose in an ancestor common to angios perms and advanced gymnosperms more than 220 million years ago. During late pollen development in angiosperms there is a switch in expression of actin s from vegetative to predominantly reproductive subclasses, perhaps to fulf il the unique functions of pollen in fertilization.