Septin Filament Formation Is Essential in Budding Yeast

Division of Biochemistry and Molecular Biology, Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA.
Developmental Cell (Impact Factor: 10.37). 04/2011; 20(4):540-9. DOI: 10.1016/j.devcel.2011.02.004
Source: PubMed

ABSTRACT Septins are GTP-binding proteins that form ordered, rod-like multimeric complexes and polymerize into filaments, but how such supramolecular structure is related to septin function was unclear. In Saccharomyces cerevisiae, four septins form an apolar hetero-octamer (Cdc11-Cdc12-Cdc3-Cdc10-Cdc10-Cdc3-Cdc12-Cdc11) that associates end-to-end to form filaments. We show that septin filament assembly displays previously unanticipated plasticity. Cells lacking Cdc10 or Cdc11 are able to divide because the now-exposed subunits (Cdc3 or Cdc12, respectively) retain an ability to homodimerize via their so-called G interface, thereby allowing for filament assembly. In such cdc10Δ and cdc11Δ cells, the remaining septins, like wild-type complexes, localize to the cortex at the bud neck and compartmentalize nonseptin factors, consistent with a diffusion barrier composed of continuous filaments in intimate contact with the plasma membrane. Conversely, Cdc10 or Cdc11 mutants that cannot self-associate, but "cap" Cdc3 or Cdc12, respectively, prevent filament formation, block cortical localization, and kill cells.

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Available from: Michael Mcmurray, Jul 25, 2014
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    • "A case in point is that septin disk enlargement by SEPT9 isoforms lacking the N-terminal extension of SEPT9(a) is not seen using the cognate AcGFP-tagged isoforms, which were nonlocalized in most cells (compare Figures 5 and 9D). Deletion of the α0 helix at the N-terminal end of the budding yeast septin CDC11 weakens homotypic CDC11–CDC11 interactions at the NC-interface of octamers, which prevents end-to-end heteromer polymerization (Bertin et al., 2008; McMurray et al., 2011). Moreover, expression of SEPT9 lacking the entire N-terminus, including the α0 helix, has been shown to disrupt septin filament formation, which supports the notion that octamers polymerize through a SEPT9–SEPT9 NC-interface (Kim et al., 2011). "
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    ABSTRACT: Septin family proteins assemble into rod-shaped heteromeric complexes that form higher-order arrangements at the cell cortex where they serve apparently conserved functions as diffusion barriers and molecular scaffolds. There are 13 confirmed septin paralogs in mammals that may be ubiquitous or tissue-specific. Septin heterooligomerization appears homology subgroup-directed, which in turn determines the subunit arrangement of six- to eight-subunit core heteromers. Here we addressed functional properties of human SEPT9 that, due to variable mRNA-splicing, exist as multiple isoforms that differ between tissues. Myeloid K562 cells express three SEPT9 isoforms, all of which have an equal propensity to heterooligomerize with SEPT7-containing hexamers to generate octameric heteromers. However, due to limiting amounts of SEPT9, K562 cells contain both hexameric and octameric heteromers. To generate cell lines with controllable hexamer to octamer ratios and that express single SEPT9 isoforms, a gene product replacement-strategy was developed. By this means we identified SEPT9 isoform-specific properties that either facilitate septin heteromer polymerization along microtubules or modulate the size-range of submembranous septin disks - a prevalent septin structure in non-adhered cells. Our findings show that the SEPT9 expression level directs the hexamer to octamer ratio, while the isoform-composition and expression level together determine higher-order arrangements of septin filaments.
    Molecular biology of the cell 09/2012; 23(21). DOI:10.1091/mbc.E12-06-0486 · 5.98 Impact Factor
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    • "Like yeast, Ashbya expresses five septins during vegetative growth, Cdc3p, Cdc10p, Cdc11p, Cdc12p, and Shs1p (Sep7p), and all are essential for ring formation (DeMay et al., 2009). In yeast, however, Shs1p is dispensable for septin ring assembly, whereas the other septins are essential for ring formation, dependent somewhat on strain background (Iwase et al., 2007; Garcia et al., 2011; McMurray et al., 2011a). "
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    ABSTRACT: Septins are conserved GTP-binding proteins that assemble into heteromeric complexes that form filaments and higher-order structures in cells. What directs filament assembly, determines the size of higher-order septin structures, and governs septin dynamics is still not well understood. We previously identified two kinases essential for septin ring assembly in the filamentous fungus Ashbya gossypii and demonstrate here that the septin Shs1p is multiphosphorylated at the C-terminus of the protein near the predicted coiled-coil domain. Expression of the nonphosphorylatable allele shs1-9A does not mimic the loss of the kinase nor does complete truncation of the Shs1p C-terminus. Surprisingly, however, loss of the C-terminus or the predicted coiled-coil domain of Shs1p generates expanded zones of septin assemblies and ectopic septin fibers, as well as aberrant cell morphology. The expanded structures form coincident with ring assembly and are heteromeric. Interestingly, while septin recruitment to convex membranes is increased, septin localization is diminished at concave membranes in these mutants. Additionally, the loss of the coiled-coil leads to increased mobility of Shs1p. These data indicate the coiled-coil of Shs1p is an important negative regulator of septin ring size and mobility, and its absence may make septin assembly sensitive to local membrane curvature.
    Molecular biology of the cell 07/2012; 23(17):3391-406. DOI:10.1091/mbc.E12-03-0207 · 5.98 Impact Factor
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    • "A recent study indicates that the survival of cells deleted for individual septin genes such as cdc10D and cdc11D is due to plasticity in septin complex and filament assembly; i.e., the remaining septin subunits are still capable of forming rod-shaped complexes that can assemble into filaments and rings, but they do so with reduced efficiency (Frazier et al. 1998; McMurray et al. 2011). Structure-based mutations that have little or no effect on septin complex formation but disrupt filament assembly invariably cause cell death, suggesting that filament assembly is essential (McMurray et al. 2011). "
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    ABSTRACT: Asymmetric cell division, which includes cell polarization and cytokinesis, is essential for generating cell diversity during development. The budding yeast Saccharomyces cerevisiae reproduces by asymmetric cell division, and has thus served as an attractive model for unraveling the general principles of eukaryotic cell polarization and cytokinesis. Polarity development requires G-protein signaling, cytoskeletal polarization, and exocytosis, whereas cytokinesis requires concerted actions of a contractile actomyosin ring and targeted membrane deposition. In this chapter, we discuss the mechanics and spatial control of polarity development and cytokinesis, emphasizing the key concepts, mechanisms, and emerging questions in the field.
    Genetics 06/2012; 191(2):347-87. DOI:10.1534/genetics.111.132886 · 4.87 Impact Factor
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