title: Kinetic partitioning during de novo septin filament assembly creates a critical G1 "window of opportunity" for mutant septin function.
abstract: Septin proteins form highly conserved cytoskeletal filaments composed of hetero-oligomers with strict subunit stoichiometry. Mutations within one hetero-oligomerization interface (the "G" interface) bias the mutant septin toward conformations that are incompatible with filament assembly, causing disease in humans and, in budding yeast cells, temperature-sensitive defects in cytokinesis. We previously found that, when the amount of other hetero-oligomerization partners is limiting, wild-type and G interface-mutant alleles of a given yeast septin "compete" along parallel but distinct folding pathways for occupancy of a limited number of positions within septin hetero-octamers. Here, we synthesize a mathematical model that outlines the requirements for this phenomenon: if a wild-type septin traverses a folding pathway that includes a single rate-limiting folding step, the acquisition by a mutant septin of additional slow folding steps creates an initially large disparity between wild-type and mutant in the cellular concentrations of oligomerization-competent monomers. When the 2 alleles are co-expressed, this kinetic disparity results in mutant exclusion from hetero-oligomers, even when the folded mutant monomer is oligomerization-competent. To test this model experimentally, we first visualize the kinetic delay in mutant oligomerization in living cells, and then narrow or widen the "window of opportunity" for mutant septin oligomerization by altering the length of the G1 phase of the yeast cell cycle, and observe the predicted exacerbation or suppression, respectively, of mutant cellular phenotypes. These findings reveal a fundamental kinetic principle governing in vivo assembly of multiprotein complexes, independent of the ability of the subunits to associate with each other.
authors.affiliation: a Department of Cell and Developmental Biology , University of Colorado Anschutz Medical Campus , Aurora , CO , USA.
authors.forename: Rachel M
authors.initials: RM
authors.lastname: Schaefer
authors.affiliation: a Department of Cell and Developmental Biology , University of Colorado Anschutz Medical Campus , Aurora , CO , USA.
authors.forename: Lydia R
authors.initials: LR
authors.lastname: Heasley
authors.affiliation: b Department of Biomedical Engineering , University of Minnesota , Minneapolis , MN , USA.
authors.forename: David J
authors.initials: DJ
authors.lastname: Odde
authors.affiliation: a Department of Cell and Developmental Biology , University of Colorado Anschutz Medical Campus , Aurora , CO , USA.
authors.forename: Michael A
authors.initials: MA
authors.lastname: McMurray
mesh_terms: D016193:G1 Phase; D007700:Kinetics; D008954:Models, Biological; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D017510:Protein Folding; D055503:Protein Multimerization; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D058112:Septins / Q000737:chemistry / Q000378:metabolism*; D013696:Temperature
chemical_list: D050505:Mutant Proteins; D029701:Saccharomyces cerevisiae Proteins; D058112:Septins
keywords: assembly; cell cycle phases; chaperones; cytoskeleton; kinetics; multiprotein complex; protein folding; septins