Formin differentially utilizes profilin isoforms to rapidly assemble actin filaments.
Cells contain multiple formin isoforms that drive the assembly of profilin-actin for diverse processes. Given that many organisms also contain several profilin isoforms, specific formin/profilin pairs might be matched to optimally stimulate actin polymerization. We utilized a combination of bulk actin polymerization and single filament total internal reflection fluorescence microscopy assays to measure the effect of different profilin isoforms on the actin assembly properties of the cytokinesis formins from fission yeast (Cdc12p) and the nematode worm (CYK-1). We discovered that Cdc12p only effectively utilizes the single fission yeast profilin isoform SpPRF. Conversely, CYK-1 prefers the essential worm cytokinesis profilin CePFN-1 to the two non-essential worm profilin isoforms (SpPRF = CePFN-1 > CePFN-2 > CePFN-3). Chimeras containing the profilin-binding formin homology 1 (FH1) domain from one formin and the barbed-end associated FH2 domain from the other formin, revealed that both the FH1 and FH2 domains help confer profilin isoform specialization. Although the Cdc12p and CYK-1 FH1 domains cannot differentiate between profilin isoforms in the absence of actin, formin FH1 domains appear to preferentially select specific isoforms of profilin-actin. Surprisingly, analysis of profilin point mutants revealed that differences in highly conserved residues in both the poly-L-proline and actin binding regions of profilin do not explain their differential utilization by formin. Therefore, rapid formin-mediated elongation of profilin-actin depends upon favorable interactions of profilin-actin with the FH1 domain as well as the barbed-end associated FH2 domain. Specific formin FH1FH2 domains are tailored to optimally utilize actin bound to particular profilin isoforms.
Neidt
Erin M
EM
Departments of Molecular Genetics and Cell Biology and Biochemistry and Molecular Biology, The University of Chicago, Chicago, Illinois 60637.
Scott
Bonnie J
BJ
Departments of Molecular Genetics and Cell Biology and Biochemistry and Molecular Biology, The University of Chicago, Chicago, Illinois 60637.
Kovar
David R
DR
Departments of Molecular Genetics and Cell Biology and Biochemistry and Molecular Biology, The University of Chicago, Chicago, Illinois 60637; Departments of Molecular Genetics and Cell Biology and Biochemistry and Molecular Biology, The University of Chicago, Chicago, Illinois 60637. Electronic address: drkovar@uchicago.edu.
D008841:Actin Cytoskeleton / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000818:Animals; D017173:Caenorhabditis elegans / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008840:Microfilament Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012568:Schizosaccharomyces / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D029742:Caenorhabditis elegans Proteins; D008840:Microfilament Proteins; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D029702:Schizosaccharomyces pombe Proteins