Septins are a group of highly-conserved GTP binding proteins found in eukaryotes. Although the mechanism of their assembly and function are not clear, they are thought to act as a component of the cytoskeleton, and thus is expected to have a large range of functions, such as in cytoskeleton rearrangement, as scaffolding proteins, and as diffusion barriers in compartmentalisation. Septin assembles first into hetero-oligomer complexes, and then may polymerize to form higher-order structures such as non-polar filaments, bundles of filaments, rings, and cage-like structures.

In 1971, Lee Hartwell published a paper^[1] entailing a set of genes that are involved in cell division, which he elucidated using site-directed mutatgensis on many genes. He discovered four genes essential for cytokinesis: cdc3, cdc10, cdc11 and cdc12. Fluorescence microscopy was then used to uncover the pattern of localisation of the gene products. It was found that all the gene products aggregate to form a ring around the neck of the division site, and thus assumed to be involved in cell septation; because of this, John Pringle (of the Pringle lab in Stanford) named these gene products 'septins'.

Septin are involved in cytokinesis, ciliogenesis, neurogenesis; polarity determination, cytoskeletal reorganization, membrane dynamics, vesicle trafficking, and exocytosis^[2]; in combating bacterial infections as part of the innate immunity. Defects in septin can lead to cancer - such as adenocarcinomas caused by increased expression of septin 9^[3], neoplasia^[4], neurodegenerative diseases, ageing, autoimmune diseases and inflammatory diseases.

All septin belongs to a supercloass of phosphate-binding loop (P-loop) NTPase.

The number of septin genes vary greatly between species. Caenorhabditis elegans have 2 septin genes, Saccharomyces cerevisiae have 7 and humans have 13 septin genes, many of which can undergo alternative splicing, often tissue specifically, to produce more variation.^[5]

The 13 septin genes in humans are SEPT1-SEPT12 and SEPT14 (SEPT13 is a pseudogene now known as SEPT7P2), and are categoriesd into four groups based on sequence homology with the septin the group is named after. The four groups are SEPT2 (includes SEPT1, SEPT2, SEPT4), SEPT3 (SEPT3, SEPT9, SEPT12), SEPT6 (SEPT6, SEPT8, SEPT10, SEPT11, SEPT14) and SEPT7 (SEPT7). Septins in the SEPT6 group lacks a Thr78 residue in the GTP-binding domain of the central core region, and thus is unable for these septins to hydrolyse GTP to GDP.a Each of the 13 human septin genes exhibit 'extensive alternate splicing'.^[6]

There are many levels of organisation and structure with septin, from a single monomer to rings. The assembly and disassembly of one level to another is triggered by signals, such as cell division checkpoints and bacterial infections.

All septin lies within the molecular range of 30-65kDa. All contains a conserved (>70% sequence similarity) central P loop-based GTP-binding domain, flanked by more flexible proline-rich N- and coiled coil C-termini. The core is made up of a small polybasic region, a large GTP-binding domain, and a septin unique element (SUE). The polybasic region binds phosphoinositides on plasma membranes, whereas SUE is a highly conserved 53-residue sequence of which the function is unclear. The length of the core genes of different septins are very similar, but there are large variations for the N- and C-termini.

Many septins can aggregate to form a rod-shaped complex. The assembly and disassembly is attributed to the hydrolysis of GTP by the GTP-binding domain, which changes the conformation of the septin monomer; this effect is likely to extend into filament formation also. The inability of septin to hydrolyse GTP (in SEPT6) leads to inability to regulate assembly and disassembly, but this can also mean stability in the complexes formed; for SEPT6, this allows it to stabilize its interaction with SEPT2, which is constitutively expressed.^[7]

There are always two copies of every septin that is in the complex, and they are arranged symmetrically, leading to a symmetrical, non-polar building block. This is in contrast to actin and microtubules, which are polar. Different organisms have different number of septins in their complex, in C. elegans, there are 2 pairs in the complex (forming a heterotetramer) and 4 pairs in S. cerevisiae (forming a heterooctamer). In humans, septin is formed as a heterohexamer (3 pairs), which can be composed of any combination of 3 from the 13 different human septin types, although the most common is the SEPT7-SEPT6-SEPT2 complex. SEPT9 (in the SEPT3 group) can be added on to SEPT7 to form an octamer, and in vivo, human septin occurs as a mixture of octamers and hexamers. In the octamer, every group of septin is represented; because there is only one member in the SEPT7 group, SEPT7 is ubiquitously expressed. Other septin groups can use different members to form a functionally seemingly-identical complex, even though the septins are different; this may be due to redundancy in some of the members of the group. This redundancy is also observed in yeast, where the lack of a member in a group is compensated by others.

When SEPT2 is overexpressed in mammalian cells, it was found to form homodimers, suggesting that the specificity of binding of different septin groups are not strict. However, when other genes (such as SEPT6 and SEPT9) were overexpressed, they remained monomers. This gives us an insight into the determining factors of septin's affinity for each other. The homodimers' and monomers' function, if any, is not known.

Another experiment where the C-terminal domain of SEPT2 is removed, SEPT2 was still able to form homodimers using the GTP-binding domain, or G interface. This suggests that the most significant binding domain is the G interface, and the N- and C-terminal domains (the NC interface) are used for specific association. This theory is contrasted when looking at the well characterised complex SEPT7-SEPT6-SEPT2-SEPT2-SEPT6-SEPT7, where the interaction alternates between using the G-interface and NC-interface; and so both types of interactions are thought to be as strong as each other, and they both exist to ensure avidity in the binding given the complexes' conformation.

The complex can associate end-to-end to form non-polar filaments, which are not as dynamic as actin and microtubules, and so is considered a more stable component of the cytoskeleton than the two, but similar to intermediate filaments.

It was found in yeast that only when the septin assembles to at least the filamentous level of organisation does it show any kind of function.

Filaments can associate laterally, with the same type of septin associating with each other, to form a bundle. A bundle can then be rearranged in various ways to give rings, meshes and cage-like structures.

Septin associates with actin, microtubules and some cell membranes, and forms the cytoskeletal structures at cell division sites, base of cilia, parts of the plasma membrane, annuli of spermatozoa, dendrites and bacterial cells.

Septin is observed to associate with actin more than any other molecules, and is thought to aid actin filament nucleation, actin-based motility and phagocytosis. When actin polymerization is inhibited by cytochalasin D, the conformation of septin changed from filaments to rings, and so it is thought that septin is involved for actin assembly.

Apart from its involvement in assembly, septin continue to assocaite with actin through actin-binding proteins. Anillin recruits septin to the actomyosin ring at the division neck to drive cell division during cytokinesis. During interphase, when anillin is localised in the nucleus, septins assocaites with non-muscle myosin II, to regulate cytoplasmic septin-actin interactions.

In contrast, when microtubules polymerization is inhibited by nocodazole, septin filaments disassemble, and so microtubules are thought to be essential for septin assembly. To reinforce this hypothesis, when microtubules are prevented from disassembling, for example using taxol, the number of septin filaments increases, and the stability of septin rings are increased.

Phospholipids are thought to aid septin assembly - septin that associates with lipid monolayers containing phosphatidylinositol-4,5-bisphosphate show a more rapid assembly than the normal self-assembly of septin. Septin has also been shown to penetrate, as well as control the shape of, cell membranes.

Whereas association with other cell structure components determine the structure, the assembly and disassembly of septin, post-translational modifications proteins are thought to be involved in the actual process. Kinases phosphorylate septin in order of it to assemble, and also determines the type of structure formed, for example, the formation of a ring at the cell division site. Sumoylation (the addition of small ubiquitin-like modifier [SUMO]) and ubiquintination are required for disassembly and degradation of septin; mutations in these sites, or mutations in the enzymes which catalyse the reactions (e.g. parkin), lead to the persistence of septin structures, and this has been linked to Parkinson's disease.

It is known that septin associates with proteins such as BORGs (binders ofRHO GTPases), which bind to SEPT6 and SEPT7 to distribute it around the cell. Depending on the type of association, septin can form different structures.

Septin is involved in the localisation of other molecules to specific places in the cell, for example recruiting SNARE proteins to the plasma membrane for vesicular fusion, or contractile proteins to the neck of the budding yeast. It can also promote interactions of the various molecules in the scaffold.

During most of the cell division process, most functional septin is in the filamentous form, having an hourglass-shaped structure. During cytokenesis, the filaments rotate 90° so they are oriented perpendicular to the axis of the neck. This allows septin to form two rings around the neck, and unlike the actomyosin ring, the septin ring's purpose is not to provide contractile force, but to recruit other proteins.

During mitotic metaphase, where condensed chromosomes align in the mid-plane of the cell so kinetochore microtubules can attach to them to be separated. For this to happen, septin align on the midplane and recruits centromere-associated protein E (CENPE), a mitotic kinesin which mediates the attachment of the microtubules to the kinetochore. CENPE also prevents aneuploidy due to the loss of a single chromosome.^[8] In cells which lack SEPT2 and SEPT7, the chromosomes did not align and thus no segregation occurred.

On the membrane, it restrain cell surface receptors, transporters and other proteins together, this ensure related enzyme products have a short distance to travel and thus lead to more rapid interactions, as well as providing the otherwise every flexible cell membrane with some rigidity. This can be seen in Xenopas laevis embryos, where the absence of septin is thought to cause blebbing.

The ability of septin to control blebbing suggests that septin is involved in regulating cell motility. During motility, septin is orientated perpendicular to the direction of travel, suggesting it provides the cell with directional information. T lymphocytes which lack septin have impaired motility, and so septin is required for the T lymphocytes to follow the chemotatic gradient.

Septin is thought to have a role as a diffusion barrier at the base of appendages (such as at the annulus of mammalian spermatozoon and at the base of a cilium) as well as in dendritic spines, and also more general areas of the plasma membrane, endoplasmic reticulum, and nuclear envelope. In cells which lack septin, compartmentalisation is lost. This is important in cytokinesis, where components from the mother cell, such as ageing factors, need to be segregated from the bud.^[9]

SEPT4-knockout mice produces spermatozoon which do not have a barrier between the anterior and posterior, making the sperm defective because the flagellum is defective, ultimately leading to a sterile male. The same knockout also produced defects in mitochondrial fission, suggesting septin work in concert with actin for fission.

In cilia, a 'ciliopathy complex' separates the ciliary membrane from the plasma membrane, and the whole complex of nine proteins are localized at the base of the cilia by SEPT2. Tubulin glutamylation, a post-translational modification of tubulin that regulates ciliary motion^[10], is also maintained by SEPT2.

Invasive bacteria have evolved mechanisms which aid them in gaining entry into the host cell, most of which involves rearrangement of the cytoskeleton, specifically actin. They can initiate this rearrangement by either interacting with the host's cell surface receptors (zippering) or by injecting signals into the cell (triggering). Once initiated, a cascade will lead to actin polymerization, using Arp2/3 as the nucleator, and cytoskeleton rearrangement, which ultimately lead to the engulfment of the bacteria. SEPT2, one of the septins found as a diffusion barrier protein, is also found to be required for assembly and recruitment around the bacteria for internalization. SEPT6 and SEPT11 seem to prevent engulfment, and their decreased expression increases the rate of bacterial internalization.

in vitro, septin are observed to form ring-like structures around bacteria; however, in vivo using Listeria monocytogenes in epithelial cells, it forms ~0.6‑μm-wide collars instead. The mechanism is unclear, but when the cells are treated with cytochalasin D, septin are not recruited at the bacterial entry site, suggesting actin polymerization is required for septin assembly.

L. monocytogenes and Shigella flexneri are two cytosolic bacteria which are able to move inside the cell by forming actin tails, "columns of clustered, branched actin fibres"

^[11]. Almost all such cytosolic bacteria make use of the host's ARP2/3 complex for polymerization. L. monocytogenes and S. flexneri activate ARP2/3 using the bacterial effector actin assembly-inducing ActA, and IcsA, respectively. Septin are recruited to these actin tails and surrounds the tail and eventually the body. Actin tail formation and septin assembly around these tails have also been observed for Listeria ivanovii, Rickettsia conorii and Mycobacterium marinum.

These septin rings around the actin tail are not thought to be involved in regulating the speed of the movement, as the RNAi-induced translation depletion of SEPT2 and SEPT9 did not change the speed of the bacteria. So this suggests a different role in septin ring formations around the actin tails.

Septin cage-like structures are found around S. flexneri; their assembly decreased actin tail formation, leading to restricted the motility as well as decreased rate of spread. As expected, the septin cage formation is preceded by actin polymerization, as cells treated with cytochalasin D did not form such structures. The septins involved are thought to be SEPT2 and SEPT9, as their decreased expression increased actin tail formation and but decreased septin cage formation. Furthermore, when SEPT2–non-muscle myosin II interactions are increased using tumour necrosis factor (TNF), few actin tails are observed, but more (~20%) are surrounded by a septin cage. These structures are shown to be stable using FRAP. Therefore, in response to motile cytosolic bacteria, septin are thought to reduce motility and dissemination of these bacteria. In other bacteria, such as L. monocytogenes, septin caging is not observed, suggesting that caging requires a specific actin polymerization pathway, or requires other factors such as neural WASP (N‑WASP). Septin cages are observed in Mycobacterium marinum, which uses the same mechanism for actin polymerization as S. flexneri.

The formation of the septin cage allows the bacterium to be targeted for autophagy, and the two process are thought to be interdependent. When the levels of SEPT2, SEPT9 or key autophagy components, such as p62, nuclear dot protein 52 kDa (NDP52), autophagyrelated 5 (ATG5), ATG6 or ATG7 are reduced, both the markers for autophagy (ubiquitin, p62 and ATG8) and septin cages are not observed. L. monocytogenes are not targeted by septin-dependent autophagy, as ActA prevents ubiquitylation of the bacteria.

"~0.6‑μm-wide cylindrical septin structures"^[11] are found in non-dividing cells of Drosophila melanogaster, and so may have functions outside bacterial infection, diffusion barrier or scaffolding (it does not co-localize with anillin.

The mechanism of septin assembly has been difficult to elucidate because

• There are many types of septin (isoforms)
• Septin are distributed differently in different cell types
• Multiple septin interact with each other, and so it is unclear about the function that each sub-unit contributes to

The mechanism of septin function is unclear because:

• Multiple septin interact with each other, but if we separate them, they may lose their functions
• Some septins are thought to be remnants of evolution, because mice with knockouts of some septin genes produced no phenotype, while knockout of other septin genes were lethal.