Some assembly required: yeast septins provide the instruction manual

Abstract

Septins are a family of conserved proteins that form hetero-oligomeric complexes that assemble into filaments. The filaments can be organized into linear arrays, coils, rings and gauzes. They serve as membrane-associated scaffolds and as barriers to demarcate local compartments, especially for the establishment of the septation site for cytokinesis. Studies in budding and fission yeast have revealed many of the protein-protein interactions that govern the formation of multi-septin complexes. GTP binding and phosphorylation direct the polymerization of filaments that is required for septin-collar assembly in budding yeast, whereas a homolog of anillin instructs timely formation of the ring of septin filaments at the medial cortex in fission yeast. These insights should aid understanding of the organization and function of the diverse septin structures in animal cells.

Figure 1. The primary structure of a typical septin and organization of septin complexes

(a) The recognizable motifs and domains in the primary structure of a septin. The GTP-binding domain (solid blue) contains all five of the signature motifs (G-boxes) that are found in other members of the super-family of GTPases [81,82]. The predicted coiled-coil sequence in the C-terminal extension (dark gray box) is preceded by a sequence that is predicted to form an α-helix (open box). (b) Proposed organization of multi-septin hetero-oligomers in eukaryotes. Models of the mitotic complexes in budding and fission yeast are adapted from Versele et al. [10] and An et al. [33], respectively. The other models are inferred from the yeast archetypes, based on sequence relatedness, similarities in the primary structure and phylogenetic analysis [5]. Some models are supported by data on individual septin-septin interactions [7,9,45]. In the human complex, Sept9 is shown as the counterpart of Cdc10 (Table 1), based on sequence similarity and phylogenetic-tree analysis [5], the lack of a CTE, and the association between Sept9 and both Sept11 (a Cdc3 ortholog) and Sept7 (a Cdc12 ortholog) [83]. The N-terminal domain of Sept9 is required for its interaction with Sept7 and Sept11, which is similar to the interaction of Cdc10 with Cdc3 and Cdc12. However, there is evidence that the C-terminal portions of Sept7 and of Sept11 are required for their interaction with Sept9 [83], unlike the interactions of Cdc3 and Cdc12 with Cdc10 [10].

Figure 2. Septin filaments from budding yeast

(a) EM images of negatively-stained preparations of filaments reconstituted from purified, recombinant septin complexes. A multimeric complex of budding yeast septins (Cdc3–Cdc12–Cdc11–Cdc10), purified from E. coli cells in which all four proteins are co-expressed, was dialysed into low-salt buffer. Previously unpublished images (left and middle panel) are courtesy of Sang-Shin Park (this laboratory) and Patricia Grob (laboratory of Eva Nogales, Univ. of California, Berkeley, USA); right panel, reproduced with permission from [10]. Scale bars, 100 nm. (b) EM images in negative-contrast of septin structures at the cell cortex of yeast spheroplasts, prepared by a rapid-freeze, deep-etch technique. Scale bars, 200 nm. Reproduced, with permission, from [13]. (c) Model of the polymerization of septin complexes into paired, linear filaments. End-to-end association of Cdc3 in one Cdc3–Cdc12–Cdc11 complex with Cdc11 in another complex yields filaments with a defined polarity; Cdc10 serves as a bridge to stabilize and pair the strands. A parallel arrangement of filaments is depicted, but an anti-parallel arrangement has not been ruled out by direct experimental evidence. Dimensions are from [8,10].

Figure 3

Schematic depiction of septin recruitment and septin-collar formation during the cell cycle of budding yeast. Recruitment of either a patch or cap of septins to the incipient bud site and its rapid conversion into a rimmed disk require the small GTPase, Cdc42. Stabilization of the disk and its transformation into a filamentous collar requires phosphorylation by two proteins kinases, Cla4 and Gin4 (and, perhaps, others), plus binding of GTP to Cdc10 and Cdc12 in the septin complex. At cytokinesis, the collar splits into two separate rings, which are disassembled after cell separation. Confocal fluorescence images (GFP–Cdc12) were kindly provided by Jeroen Dobbelaere (ETH, Zürich, Switzerland).