Null mutants of the Neurospora actin-related protein 1 pointed-end complexshow distinct phenotypes

Dynactin is a multisubunit complex that regulates the activities of cytopla smic dynein, a microtubule-associated motor. Actin-related protein 1 (Arp1) is the most abundant subunit of dynactin, and it forms a short filament to which additional subunits associate. An Arp1 filament pointed-end-binding subcomplex has been identified that consists of p62, p25, p27, and Arp11 su bunits. The functional roles of these subunits have not been determined. Re cently, we reported the cloning of an apparent homologue of mammalian Arp11 from the filamentous fungus Neurospora crassa. Here, we report that N. cra ssa ro-2 and ro-12 genes encode the respective p62 and p25 subunits of the pointed-end complex. Characterization of Delta ro-2, Delta ro-7, and Delta ro-12 mutants reveals that each has a distinct phenotype. All three mutants have reduced in vivo vesicle trafficking and have defects in vacuole distr ibution. We showed previously that in vivo dynactin function is required fo r high-level dynein ATPase activity, and we find that all three mutants hav e low dynein ATPase activity. Surprisingly, Delta ro-12 differs from Delta ro-2 and Delta ro-7 and other previously characterized dynein/dynactin muta nts in that it has normal nuclear distribution. Each of the mutants shows a distinct dynein/dynactin localization pattern. All three mutants also show stronger dynein/dynactin-membrane interaction relative to wild type, sugge sting that the Delta rp1. pointed-end complex may regulate interaction of d ynactin with membranous cargoes.