The Arp2/3 complex nucleates actin arrays during zygote polarity establishment and growth

Previous work has demonstrated that dynamic actin arrays are important for axis establishment and polar growth in the fucoid zygote, Silvetia compressa. Transitions between these arrays are mediated by depolymerization of an existing array and polymerization of a new array. To begin to understand how polymerization of new arrays might be regulated, we investigated the role of the highly conserved, actin-nucleating, Actin-related protein 2/3 (Arp2/3) complex. Arp2, a subunit of the complex, was cloned and peptide antibodies were raised to the C-terminal domain. In immunolocalization studies of polarizing zygotes, actin and Arp2 colocalized around the nucleus and in a patch at the rhizoid pole. In germinated zygotes, a cone of Arp2 and actin extended from the nucleus to the subapex. Within the rhizoid tip, three structural zones were observed in the majority of zygotes: the extreme apex was devoid of label, the subapex was enriched for Arp2, and further back both actin and Arp2 were present. This zonation suggests that actin nucleation occurs at the leading edge of the cone, in the Arp2-enriched region. In two sets of experiments, we showed that tip zonation is important for growth. First, pharmacological treatments that disrupted Arp2/actin zonation arrested tip growth. Second, changes in the direction of tip growth during negative phototropism were preceded by a reorientation of the zonation in accordance with the new growth direction. This work represents the first investigation of Arp2/3 complex localization in tip-growing algal cells.