Mechanism and biological role of profilin-Srv2/CAP interaction.
Profilin and cyclase-associated protein (CAP, known in yeast as Srv2) are ubiquitous and abundant actin monomer-binding proteins. Profilin catalyses the nucleotide exchange on actin monomers and promotes their addition to filament barbed ends. Srv2/CAP recycles newly depolymerized actin monomers from ADF/cofilin for subsequent rounds of polymerization. Srv2/CAP also harbors two proline-rich motifs and has been suggested to interact with profilin. However, the mechanism and biological role of the possible profilin-Srv2/CAP interaction has not been investigated. Here, we show that Saccharomyces cerevisiae Srv2 and profilin interact directly (K(D) approximately 1.3 microM) and demonstrate that a specific proline-rich motif in Srv2 mediates this interaction in vitro and in vivo. ADP-actin monomers and profilin do not interfere with each other's binding to Srv2, suggesting that these three proteins can form a ternary complex. Genetic and cell biological analyses on an Srv2 allele (srv2-201) defective in binding profilin reveals that a direct interaction with profilin is not essential for Srv2 cellular function. However, srv2-201 causes a moderate increase in cell size and partially suppresses the cell growth and actin organization defects of an actin binding mutant profilin (pfy1-4). Together these data suggest that Srv2 is an important physiological interaction partner of profilin.
Bertling
Enni
E
Institute of Biotechnology, University of Helsinki, 00014 Helsinki, Finland.
Quintero-Monzon
Omar
O
Mattila
Pieta K
PK
Goode
Bruce L
BL
Lappalainen
Pekka
P
D000199:Actins / Q000032:analysis / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing; D000483:Alleles; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D048429:Cell Size; D003598:Cytoskeletal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D047468:Immunoprecipitation; D007700:Kinetics; D009154:Mutation; D051304:Profilins / Q000235:genetics / Q000378:metabolism*; D011392:Proline / Q000737:chemistry; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000166:cytology / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D003598:Cytoskeletal Proteins; D051304:Profilins; C073751:SRV2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D011392:Proline