Heterotypic Coiled-Coil Formation is Essential for the Correct Assembly of the Septin Heterofilament

Sala, F.; Valadares, Napoleão Fonseca; Macêdo, Joci Neuby Alves; Borges, Júlio César; Garratt, R.C.

doi:10.1016/j.bpj.2016.10.032

Cited by 38 publications

(43 citation statements)

References 39 publications

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“…For example, human septins of the SEPT6 and SEPT7 groups present a long C-terminal domain with more than 100 residues, while this region presents only an intermediate length in members of the SEPT2 group and is smallest of all in the SEPT3-like septins. As mentioned above, with the exception of the SEPT3 group, the sequences of the C-terminal domains are predicted to form coiled coils, and biophysical studies have shown that the C-termini of septins of the SEPT6 group associate with high affinity with those of SEPT7, forming stable hetero-dimeric coiled coils (Marques et al 2012;Sala et al 2016). The pairing of the coiled coil regions of SEPT6-like septins with SEPT7 is consistent with the predicted number of heptad repeats, which is greater than that expected for SEPT2 and its group members.…”

Section: A Brief Summarymentioning

confidence: 95%

“…Two types of generic interface, G and NC, alternate along the filament. These can be further subdivided into NC, NC het and NC hom for the NC-interfaces and G DD and G DT for the G-interfaces (further explanation is given in the text) the C-domain may play a dominant role in selective assembly (Marques et al 2012;Sala et al 2016).…”

Section: Recollection On Septin Structural Studies Conducted During Tmentioning

confidence: 99%

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Septin structure and filament assembly

et al. 2017

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Septins are able to polymerize into long apolar filaments and have long been considered to be a component of the cytoskeleton alongside intermediate filaments (which are also apolar in nature), microtubules and actin filaments (which are not). Their central guanosine triphosphate (GTP)-binding domain, which is essential for stabilizing the filament itself, is flanked by N- and C-terminal domains for which no direct structural information is yet available. In most cases, physiological filaments are built from a number of different septin monomers, and in the case of mammalian septins this is most commonly either three or four. Comprehending the structural basis for the spontaneous assembly of such filaments requires a deeper understanding of the interfaces between individual GTP-binding domains than is currently available. Nevertheless, in this review we will summarize the considerable progress which has been made over the course of the last 10 years. We will provide a brief description of each structure determined to date and comment on how it has added to the body of knowledge which is rapidly growing. Rather than simply repeat data which have already been described in the literature, as far as is possible we will try to take advantage of the full set of information now available (mostly derived from human septins) and draw the reader's attention to some of the details of the structures themselves and the filaments they form which have not be commented on previously. An additional aim is to clarify some misconceptions.

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Section: A Brief Summarymentioning

confidence: 95%

Section: Recollection On Septin Structural Studies Conducted During Tmentioning

confidence: 99%

Septin structure and filament assembly

et al. 2017

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“…Each septin monomer participates in two interfaces which alternate along the filament: the NC interface, involving the N and C terminal helices of the G domains; and the G interface, including the region directly involved in guanine‐nucleotide‐binding (Sirajuddin et al, ; Valadares et al, ). The remaining domains (particularly the coiled coil region of the C‐domain) also contribute to the affinity and specificity of the NC interface (Barth, Schoeffler, & Alber, ; de Marques et al, ; Sala, Valadares, Macedo, Borges, & Garratt, ; Versele et al, ).…”

Section: Introductionmentioning

confidence: 99%

A revised order of subunits in mammalian septin complexes

Mendonça¹,

Macêdo

Guimarães³

et al. 2019

Cytoskeleton

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Septins are GTP binding proteins considered to be novel components of the cytoskeleton. They polymerize into filaments based on hexameric or octameric core particles in which two copies of either three or four different septins, respectively, assemble into a specific sequence. Viable combinations of the 13 human septins are believed to obey substitution rules in which the different septins involved must come from distinct subgroups. The hexameric assembly, for example, has been reported to be SEPT7–SEPT6–SEPT2–SEPT2–SEPT6–SEPT7. Here, we have replaced SEPT2 by SEPT5 according to the substitution rules and used transmission electron microscopy to demonstrate that the resulting recombinant complex assembles into hexameric particles which are inverted with respect that predicted previously. MBP‐SEPT5 constructs and immunostaining show that SEPT5 occupies the terminal positions of the hexamer. We further show that this is also true for the assembly including SEPT2, in direct contradiction with that reported previously. Consequently, both complexes expose an NC interface, as reported for yeast, which we show to be more susceptible to high salt concentrations. The correct assembly for the canonical combination of septins 2‐6‐7 is therefore established to be SEPT2–SEPT6–SEPT7–SEPT7–SEPT6–SEPT2, implying the need for revision of the mechanisms involved in filament assembly.

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“…Each septin monomer participates in two interfaces which alternate along the filament: the NC interface, involving the N and C terminal helices of the G domains; and the G interface, including the region directly involved in guanine-nucleotide-binding [3,13]. The remaining domains (particularly the coiled coil region of the C-domain) also contribute to the affinity and specificity of the NC interface [15,16]. Sirajuddin et al, 2007, using negative-stain electron microscopy and employing a version of SEPT2 fused to MBP, proposed that SEPT2 occupies the central position of the core complex and SEPT7 lies at its extremities, leading to the following arrangement for the hexamer: SEPT7-SEPT6-SEPT2-SEPT2-SEPT6-SEPT7 (which, for convenience, we will abbreviate to 7-6-2-2-6-7).…”

Section: Introductionmentioning

confidence: 99%

Repositioning septins within the core particle

Mendonça

Macêdo

Itri³

et al. 2019

Preprint

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Abbreviations (Define non-standard or uncommon abbreviations). AbstractSeptins are GTP binding proteins considered to be a novel component of the cytoskeleton. They polymerize into filaments based on hetero-oligomeric core particles which, in humans, are either hexamers or octamers composed of two copies each of either three or four different septins from the 13 available. Not all combinations are possible as it is believed that these must obey substitution rules which determine that different septins must be derived from four distinct and well-established sub-groups. Here, we have purified and characterized one such combinations, SEPT5-SEPT6-SEPT7, and used TEM to derive the first structural information concerning its assembly. The full complex was purified using an affinity tag attached to only one of its components (SEPT7) and was able to bind to and perturb lipid bilayers. Although the complex assembled into elongated hexameric particles, the position of SEPT5 was incompatible with that predicted by the reported structure of SEPT2-SEPT6-SEPT7 based on the substitution rules. MBP-fusion constructs for SEPT5 and SEPT2 and immuno-staining clearly show that these septins occupy the terminal positions of the SEPT5-SEPT6-SEPT7 and SEPT2-SEPT6-SEPT7 hexamers, respectively. In so doing they expose a so-called NC interface which we show to be more susceptible to perturbation at high salt concentrations. Our results show that the true structure of the hexamer is inverted with respect to that described previously and, as such, is more compatible with that reported for yeast. Taken together, our results suggest that the mechanisms involved in spontaneous self-assembly of septin core particles and their filaments deserve further reflection.

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Heterotypic Coiled-Coil Formation is Essential for the Correct Assembly of the Septin Heterofilament

Cited by 38 publications

References 39 publications

Septin structure and filament assembly

Septin structure and filament assembly

A revised order of subunits in mammalian septin complexes

Repositioning septins within the core particle

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