Abstract
Human septins 3, 9 and 12 are the only members of a specific subgroup of septins that display several unusual features, including the absence of a C-terminal coiled coil. This particular subgroup (the SEPT3 septins) are present in rod-like octameric protofilaments but are lacking in similar hexameric assemblies, which only contain representatives of the three remaining subgroups. Both hexamers and octamers can self-assemble into mixed filaments by end-to-end association, implying that the SEPT3 septins may facilitate polymerization but not necessarily function. These filaments frequently associate into higher order complexes which associate with biological membranes, triggering a wide range of cellular events. In the present work, a complete compendium of crystal structures for the GTP-binding domains of all of the SEPT3 subgroup members when bound to either GDP or to a GTP analogue is provided. The structures reveal a unique degree of plasticity at one of the filamentous interfaces (dubbed NC). Specifically, structures of the GDP and GTPγS complexes of SEPT9 reveal a squeezing mechanism at the NC interface which would expel a polybasic region from its binding site and render it free to interact with negatively charged membranes. On the other hand, a polyacidic region associated with helix α5', the orientation of which is particular to this subgroup, provides a safe haven for the polybasic region when retracted within the interface. Together, these results suggest a mechanism which couples GTP binding and hydrolysis to membrane association and implies a unique role for the SEPT3 subgroup in this process. These observations can be accounted for by constellations of specific amino-acid residues that are found only in this subgroup and by the absence of the C-terminal coiled coil. Such conclusions can only be reached owing to the completeness of the structural studies presented here.
Highlights
Septins are GTP-binding proteins that are involved in important cellular processes such as cytokinesis, membrane trafficking and microtubule dynamics
IUCrJ (2020). 7, 462–479 research papers al., 2008). Many of these biological functions are dependent on the intrinsic ability of septins to spontaneously polymerize into filaments, which subsequently assemble into higher order structures such as rings and networks that are capable of membrane association
Most septins are characterized by three distinct structural domains: a variable N-terminal domain including a polybasic region that is capable of interacting with specific membrane components (Casamayor & Snyder, 2003; Zhang et al, 1999), a central GTP-binding domain (G domain) including the socalled septin unique element (SUE; Versele & Thorner, 2005), and a C-terminal domain that normally includes heptad repeats characteristic of coiled coils (Pan et al, 2007; Versele et al, 2004)
Summary
Septins are GTP-binding proteins that are involved in important cellular processes such as cytokinesis, membrane trafficking and microtubule dynamics. It has recently been shown that the order of septins within the core complexes is most likely to be SEPT2–SEPT6–SEPT7–SEPT7–SEPT6– SEPT2 for hexamers and SEPT2–SEPT6–SEPT7–SEPT9– SEPT9–SEPT7–SEPT6–SEPT2 for octamers (Mendonca et al, 2019; Soroor et al, 2019) These can associate end to end to form mixed apolar filaments (Soroor et al, 2019) that present two alternating interfaces known as G and NC (Sirajuddin et al, 2007; Valadares et al, 2017). Their conserved structural features shed light on Kinoshita’s proposal for substitutability between septins from the same subgroup within heterocomplexes
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