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Abstract Morphological complexity and plasticity are hallmarks of polyextremotolerant fungi. Septins are conserved cytoskeletal proteins and key contributors to cell polarity and morphogenesis. They sense membrane curvature, coordinate cell division, and influence diffusion at the plasma membrane. Four septins homologs are conserved from yeasts to humans, the two systems in which septins have been studied most extensively. But there is also a fifth family of septin proteins that remain biochemically mysterious. Members of this family, known as Group 5 septins, appear in the genomes of filamentous fungi, and thus have been understudied due to their absence from ascomycete yeasts.Knufia petricolais an emerging model polyextremotolerant black fungus that can serve as a model system for understudied Group 5 septins. We have recombinantly expressed and biochemically characterizedKpAspE, a Group 5 septin fromK. petricola, demonstrating that this septin––by itselfin vitro–– recapitulates many of the functions of canonical septin hetero-octamers.KpAspE is an active GTPase that forms diverse homo-oligomers, senses membrane curvature, and interacts with the terminal subunit of canonical septin hetero-octamers. These findings raise the possibility that Group 5 septins govern the higher order structures formed by canonical septins, which inK. petricolacells form extended filaments. These findings provide insight into how septin hetero-oligomers evolved from ancient homomers and raise the possibility that Group 5 septins govern the higher order structures formed by canonical septins. Significance StatementSeptins are understudied cytoskeletal proteins. Here, we biochemically characterizedKpAspE, a novel Group 5 septin from a polyextremotolerant black fungus.KpAspE in isolation recapitulates many properties of canonical septin hetero-octamersin vitroand interacts with the Cdc11, the terminal subunit of those octamers.These findings provide insight into how ancient septins may have evolved and diversified from homopolymers and suggest that many of the septin functions were present in the ancestral protein.
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Septins regulate border cell surface geometry, shape, and motility downstream of Rho in Drosophila
Septins self-assemble into polymers that bind and deform membranes in vitro and regulate diverse cell behaviors in vivo. How their in vitro properties relate to their in vivo functions is under active investigation. Here, we uncover requirements for septins in detachment and motility of border cell clusters in the Drosophila ovary. Septins and myosin colocalize dynamically at the cluster periphery and share phenotypes but, surprisingly, do not impact each other. Instead, Rho independently regulates myosin activity and septin localization. Active Rho recruits septins to membranes, whereas inactive Rho sequesters septins in the cytoplasm. Mathematical analyses identify how manipulating septin expression levels alters cluster surface texture and shape. This study shows that the level of septin expression differentially regulates surface properties at different scales. This work suggests that downstream of Rho, septins tune surface deformability while myosin controls contractility, the combination of which governs cluster shape and movement.
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- PAR ID:
- 10516931
- Publisher / Repository:
- Elsevier Inc.
- Date Published:
- Journal Name:
- Developmental Cell
- Volume:
- 58
- Issue:
- 15
- ISSN:
- 1534-5807
- Page Range / eLocation ID:
- 1399 to 1413.e5
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Momany, Michelle (Ed.)Morphological complexity and plasticity are hallmarks of polyextremotolerant fungi. Septins are conserved cytoskeletal proteins and key contributors to cell polarity and morphogenesis. They sense membrane curvature, coordinate cell division, and influence diffusion at the plasma membrane. Four septin homologues are conserved from yeasts to humans, the systems in which septins have been most studied. But there is also a fifth family of opisthokont septins that remain biochemically mysterious. Members of this family, Group 5 septins, appear in the genomes of filamentous fungi, but are understudied due to their absence from ascomycete yeasts. Knufia petricola is an emerging model polyextremotolerant black fungus that can also serve as a model system for Group 5 septins. We have recombinantly expressed and biochemically characterized KpAspE, a Group 5 septin from K. petricola. This septin––by itself in vitro––recapitulates many functions of canonical septin hetero-octamers. KpAspE is an active GTPase that forms diverse homo-oligomers, binds shallow membrane curvatures, and interacts with the terminal subunit of canonical septin hetero-octamers. These findings raise the possibility that Group 5 septins govern the higher-order structures formed by canonical septins, which in K. petricola cells form extended filaments, and provide insight into how septin hetero-oligomers evolved from ancient homomers.more » « less
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ABSTRACT Cells employ cytoskeletal polymers to move, divide, and pass information inside and outside of the cell. Previous work on eukaryotic cytoskeletal elements such as actin, microtubules, and intermediate filaments investigating the mechanisms of polymerization have been critical to understand how cells control the assembly of the cytoskeleton. Most biophysical analyses have considered cooperative versus isodesmic modes of polymerization; this framework is useful for specifying functions of regulatory proteins that control nucleation and understanding how cells regulate elongation in time and space. The septins are considered a fourth component of the eukaryotic cytoskeleton, but they are poorly understood in many ways despite their conserved roles in membrane dynamics, cytokinesis, and cell shape, and in their links to a myriad of human diseases. Because septin function is intimately linked to their assembled state, we set out to investigate the mechanisms by which septin polymers elongate under different conditions. We used simulations,in vitroreconstitution of purified septin complexes, and quantitative microscopy to directly interrogate septin polymerization behaviors in solution and on synthetic lipid bilayers of different geometries. We first used reactive Brownian dynamics simulations to determine if the presence of a membrane induces cooperativity to septin polymerization. We then used fluorescence correlation spectroscopy (FCS) to assess septins’ ability to form filaments in solution at different salt conditions. Finally, we investigated septin membrane adsorption and polymerization on planar and curved supported lipid bilayers. Septins clearly show signs of salt-dependent cooperative assembly in solution, but cooperativity is limited by binding a membrane. Thus, septin assembly is dramatically influenced by extrinsic conditions and substrate properties and can show properties of both isodesmic and cooperative polymers. This versatility in assembly modes may explain the extensive array of assembly types, functions, and subcellular locations in which septins act. SIGNIFICANCEThe septin cytoskeleton plays conserved and essential roles in cell division, membrane remodeling, and intracellular signaling with links to varied human diseases. Unlike actin and microtubules, whose polymerization dynamics have been extensively characterized, the molecular details of septin polymerization remain poorly understood. Here, we investigate the mode of septin polymerization through the lens of isodesmic and cooperative polymer assembly models in solution, on planar and curved supported membranes, and under different ionic conditions. Our findings show that the mechanisms of septin assembly are highly sensitive to ionic conditions, membrane geometry, and protein concentrations. Notably, assembly can show either cooperative or isodesmic properties depending on context, thereby revealing unexpected plasticity.more » « less
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Summary Septins, a conserved family of filament-forming proteins, contribute to eukaryotic cell division, polarity, and membrane trafficking. Septins scaffold other proteins to cellular membranes, but it is not fully understood how septins associate with membranes. We identified and characterized an isoform ofCaenorhabditis elegansseptin UNC-61 that was predicted to contain a transmembrane domain (TMD). The TMD isoform is expressed in a subset of tissues where the known septins were known to act, and TMD function was required for tissue integrity of the egg-laying apparatus. We found predicted TMD-containing septins across much of opisthokont phylogeny and demonstrated that the TMD-containing sequence of a primate TMD-septin is sufficient for localization to cellular membranes. Together, our findings reveal a novel mechanism of septin-membrane association with profound implications for septin dynamics and regulation.more » « less
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A gene duplication of a septin reveals a developmentally regulated filament length control mechanismSeptins are a family of conserved filament-forming proteins that function in multiple cellular processes. The number of septin genes within an organism varies, and higher eukaryotes express many septin isoforms due to alternative splicing. It is unclear if different combinations of septin proteins in complex alter the polymers’ biophysical properties. We report that a duplication event within the CDC11 locus in Ashbya gossypii gave rise to two similar but distinct Cdc11 proteins: Cdc11a and Cdc1b. CDC11b transcription is developmentally regulated, producing different amounts of Cdc11a- and Cdc11b-complexes in the lifecycle of Ashbya gossypii. Deletion of either gene results in distinct cell polarity defects, suggesting non-overlapping functions. Cdc11a and Cdc11b complexes have differences in filament length and membrane-binding ability. Thus, septin subunit composition has functional consequences on filament properties and cell morphogenesis. Small sequence differences elicit distinct biophysical properties and cell functions of septins, illuminating how gene duplication could be a driving force for septin gene expansions seen throughout the tree of life.more » « less
- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1016/j.devcel.2023.05.017
