Genetic dissection of the budding yeast Arp2/3 complex: A comparison of the in vivo and structural roles of individual subunits

In previous work, we identified the yeast Arp2/3 complex, which localizes t o cortical actin patches and is required for their motility and integrity i n vivo. This complex contains proteins homologous to each subunit of the Ac anthamoeba and human Arp2/3 complex except for a 40-kDa subunit (p40), whic h,vas missing from the purified yeast complex. Here, we demonstrate by usin g immunoprecipitation and gel-filtration analysis that Arc40p, the homolog of p40 identified from the yeast genome database, associates with the yeast Arp2/3 complex. We have carried out gene disruptions of each subunit of th e yeast Arp2/3 complex to study each subunits role in the function of the c omplex, Surprisingly,,ve find that only ARC40 is fully essential for cell v iability. Strains lacking each of the other subunits exhibit varying degree s of defects in cell growth and viability and in assembly and polarization of cortical actin patches. We have also examined each subunit's role in mai ntaining the structural integrity of the Arp2/3 complex, Arp2p Arp3p, and A rc40p fall into the monomer pool in Delta arc19 and Delta arc35 cells, sugg esting that Arc19p and Arc35p are the central scaffolding components of the complex, Arp2p and Arp3p do not have major roles in maintaining complex in tegrity, and Arc15p is required for association of Arp2p and Arc40p. but no t other subunits, with the complex These results provide evidence that each subunit contributes differently to the assembly and function of the Arp2/3 complex.