The complex containing actin-related proteins Arp2 and Arp3 is required for the motility and integrity of yeast actin patches.
BACKGROUND
Structural modeling and biochemical experiments in vitro have implicated a multi-protein complex containing two actin-related proteins, Arp2 and Arp3, as a potential actin-filament nucleation factor. This 'Arp2/3 complex' has been identified in Acanthamoeba and human cells and has been shown to localize to regions involved in actin-based motility, such as the leading edge of moving cells and the 'tail' of actin that forms behind the intracellular pathogen Listeria. The function of this complex in vivo has not been characterized, however, and the sequences of the non-actin-related subunits remain to be determined.


RESULTS
An Arp3 homologue from the budding yeast Saccharomyces cerevisiae was found to localize to cortical actin patches, highly motile structures that concentrate at sites of polarized growth during the yeast cell cycle. A conditional arp3 mutant allele inhibited cortical actin motility at the restrictive temperature and eventually disrupted actin patches. Most Arp3 protein is found in a multi-protein complex; we purified this complex and determined the sequences of each of the protein subunits using a high-accuracy mass peptide-mapping technique. The proteins found in the complex are similar to those in the Acanthamoeba and human Arp2/3 complexes except that the yeast complex lacks a 40 kDa subunit, which is therefore not required for the structural integrity of the complex.


CONCLUSIONS
The Arp2/3 protein complex is conserved from yeast to man, and in yeast the complex is required in vivo for the motility and integrity of cortical actin patches. We hypothesize that these patches may move by a Listeria-like mechanism driven by actin polymerization.
Winter
D
D
Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA.
Podtelejnikov
A V
AV
Mann
M
M
Li
R
R
D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000032:analysis / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D000595:Amino Acid Sequence; D003598:Cytoskeletal Proteins; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D016296:Mutagenesis; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000502:physiology*
D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; D003598:Cytoskeletal Proteins; D005656:Fungal Proteins