The MoLfa1 Protein Regulates Fungal Development and Septin Ring Formation in Magnaporthe oryzae.

Septins play a key regulatory role in cell division, cytokinesis, and cell polar growth of the rice blast fungus (Magnaporthe oryzae). We found that the organization of the septin ring, which is essential for appressorium-mediated infection in M. oryzae, requires long-chain fatty acids (LCFAs), which act as mediators of septin organization at membrane interfaces. However, it is unclear how septin ring formation and LCFAs regulate the pathogenicity of the rice blast fungus. In this study, a novel protein was named MoLfa1 because of its role in LCFAs utilization. MoLfa1 affects the utilization of LCFAs, lipid metabolism, and the formation of the septin ring by binding with phosphatidylinositol phosphates (PIPs), thereby participating in the construction of penetration pegs of M. oryzae. In addition, MoLfa1 is localized in the endoplasmic reticulum (ER) and interacts with the ER-related protein MoMip11 to affect the phosphorylation level of Mps1. (Mps1 is the core protein in the MPS1-MAPK pathway.) In conclusion, MoLfa1 affects conidia morphology, appressorium formation, lipid metabolism, LCFAs utilization, septin ring formation, and the Mps1-MAPK pathway of M. oryzae, influencing pathogenicity.

Exocytic plasma membrane flows remodel endoplasmic reticulum-plasma membrane tethering for septin collar assembly.

Endoplasmic reticulum (ER)-plasma membrane (PM) tethering is crucial for the non-vesicular lipid transport between the ER membrane and the PM. However, the PM-associated ER can impede the PM binding of cytoskeletons and other organelles. It is poorly understood how the competition between the ER and cytoskeletons/organelles on the PM is resolved. Here, we show that, upon septin collar assembly, ER-PM tethering proteins are excluded from the yeast bud sites, and the PM-associated ER is locally detached from the PM. Our results suggest that PM flows by polarized exocytosis extrude PM proteins, including ER-PM tethering proteins, from the bud sites. When the reorganization of the ER-PM tethering was inhibited by exocytosis repression, septin localization was restricted to the PM sites poor in ER-PM tethering proteins. This study proposes machinery reconciling ER-septin competition on the PM, providing mechanistic insights into the spatial organization of PM-associated organelles and cytoskeletons.

Unearthing the role of septins in viral infections.

Septin proteins are a subfamily of closely related GTP-binding proteins conserved in all species except for higher plants and perform essential biological processes. Septins self-assemble into heptameric or octameric complexes and form higher-order structures such as filaments, rings, or gauzes by end-to-end binding. Their close association with cell membrane components makes them central in regulating critical cellular processes. Due to their organisation and properties, septins function as diffusion barriers and are integral in providing scaffolding to support the membrane's curvature and stability of its components. Septins are also involved in vesicle transport and exocytosis through the plasma membrane by co-localising with exocyst protein complexes. Recently, there have been emerging reports of several human and animal diseases linked to septins and abnormalities in their functions. Most of our understanding of the significance of septins during microbial diseases mainly pertains to their roles in bacterial infections but not viruses. This present review focuses on the known roles of septins in host-viral interactions as detailed by various studies.

Chemical Proteomics Reveals Nε-Fatty-Acylation of Septins by Rho Inactivation Domain (RID) of the Vibrio MARTX Toxin to Alter Septin Localization and Organization.

Vibrio species, the Gram-negative bacterial pathogens causing cholera and sepsis, produce multiple secreted virulence factors for infection and pathogenesis. Among these is the multifunctional-autoprocessing repeats-in-toxin (MARTX) toxin that releases several critical effector domains with distinct functions inside eukaryotic host cells. One such effector domain, the Rho inactivation domain (RID), has been discovered to catalyze long-chain Nε-fatty-acylation on lysine residues of Rho GTPases, causing inactivation of Rho GTPases and disruption of the host actin cytoskeleton. However, whether RID modifies other host proteins to exert additional functions remains to be determined. Herein, we describe the integration of bioorthogonal chemical labeling and quantitative proteomics to globally profile the target proteins modified by RID in living cells. More than 246 proteins are identified as new RID substrates, including many involved in GTPase regulation, cytoskeletal organization, and cell division. We demonstrate that RID extensively Nε-fatty-acylates septin proteins, the fourth cytoskeletal component of mammalian cells with important roles in diverse cellular processes. While affinity purification and mass spectrometry analysis show that RID-mediated Nε-fatty-acylation does not affect septin-septin interactions, this modification increases the membrane association of septins and confers localization to detergent-resistant membrane rafts. As a result, the filamentous assembly and organization of septins are disrupted by RID-mediated Nε-fatty-acylation, further contributing to cytoskeletal and mitotic defects that phenocopy the effects of septin depletion. Overall, our work greatly expands the substrate scope and function of RID and demonstrates the role of RID-mediated Nε-fatty-acylation in manipulating septin localization and organization.

Palmitoylation is required for Sept8-204 and Sept5 to form vesicle-like structure and colocalize with synaptophysin.

Sept8 is a vesicle associated protein and there are two typical transcriptional variants (Sept8-204 and Sept8-201) expressed in mice brain. Interestingly, the coexpression of Sept8-204/Sept5 induces the formation of small sized vesicle-like structure, while that of the Sept8-201/Sept5 produces large puncta. Sept8 is previously shown to be palmitoylated. Here it was further revealed that protein palmitoylation is required for Sept8-204/Sept5 to maintain small sized vesicle-like structure and colocalize with synaptophysin, since either the expression of nonpalmitoylated Sept8-204 mutant (Sept8-204-3CA) or inhibiting Sept8-204 palmitoylation by 2-BP with Sept5 produces large puncta, which barely colocalizes with synaptophysin (SYP). Moreover, it was shown that the dynamic palmitoylation of Sept8-204 is controlled by ZDHHC17 and PPT1, loss of ZDHHC17 decreases Sept8-204 palmitoylation and induces large puncta, while loss of PPT1 increases Sept8-204 palmitoylation and induces small sized vesicle-like structure. Together, these findings suggest that palmitoylation is essential for the maintenance of the small sized vesicle-like structure for Sept8-204/Sept5, and may hint their important roles in synaptic functions.

Septins provide microenvironment sensing and cortical actomyosin partitioning in motile amoeboid T lymphocytes.

The all-terrain motility of lymphocytes in tissues and tissue-like gels is best described as amoeboid motility. For amoeboid motility, lymphocytes do not require specific biochemical or structural modifications to the surrounding extracellular matrix. Instead, they rely on changing shape and steric interactions with the microenvironment. However, the exact mechanism of amoeboid motility remains elusive. Here, we report that septins participate in amoeboid motility of T cells, enabling the formation of F-actin and α-actinin-rich cortical rings at the sites of cell cortex-indenting collisions with the extracellular matrix. Cortical rings compartmentalize cells into chains of spherical segments that are spatially conformed to the available lumens, forming transient "hourglass"-shaped steric locks onto the surrounding collagen fibers. The steric lock facilitates pressure-driven peristaltic propulsion of cytosolic content by individually contracting cell segments. Our results suggest that septins provide microenvironment-guided partitioning of actomyosin contractility and steric pivots required for amoeboid motility of T cells in tissue-like microenvironments.

PCP and Septins govern the polarized organization of the actin cytoskeleton during convergent extension.

Convergent extension (CE) requires the coordinated action of the planar cell polarity (PCP) proteins1,2 and the actin cytoskeleton,3,4,5,6 but this relationship remains incompletely understood. For example, PCP signaling orients actomyosin contractions, yet actomyosin is also required for the polarized localization of PCP proteins.7,8 Moreover, the actin-regulating Septins play key roles in actin organization9 and are implicated in PCP and CE in frogs, mice, and fish5,6,10,11,12 but execute only a subset of PCP-dependent cell behaviors. Septin loss recapitulates the severe tissue-level CE defects seen after core PCP disruption yet leaves overt cell polarity intact.5 Together, these results highlight the general fact that cell movement requires coordinated action by distinct but integrated actin populations, such as lamella and lamellipodia in migrating cells13 or medial and junctional actin populations in cells engaged in apical constriction.14,15 In the context of Xenopus mesoderm CE, three such actin populations are important, a superficial meshwork known as the "node-and-cable" system,4,16,17,18 a contractile network at deep cell-cell junctions,6,19 and mediolaterally oriented actin-rich protrusions, which are present both superficially and deeply.4,19,20,21 Here, we exploited the amenability of the uniquely "two-dimensional" node and cable system to probe the relationship between PCP proteins, Septins, and the polarization of this actin network. We find that the PCP proteins Vangl2 and Prickle2 and Septins co-localize at nodes, and that the node and cable system displays a cryptic, PCP- and Septin-dependent anteroposterior (AP) polarity in its organization and dynamics.

External quality assessment for detection of colorectal cancer by Septin9 DNA methylation in clinical laboratories.

BACKGROUND AND AIMS: The incidence and mortality rate of colorectal cancer (CRC) are increasing worldwide. Septin9 methylated (mSEPT9) DNA in circulation can be used as a non-invasive detection method to assist in the early diagnosis of CRC; however, the detection methods and procedures are complicated. This study aimed to evaluate the ability of clinical laboratories to detect Septin9 methylation in plasma cell-free DNA (cfDNA). MATERIALS AND METHODS: We prepared a sample panel consisting of positive and negative Septin9 methylation cells and CRC cells. Three positive samples with different methylation levels, one negative sample and one duplicate sample, two samples containing interference, three different CRC cell samples, and a fictitious case report were included. The panel was distributed to 59 laboratories for mSEPT9 analysis, result comparison, and scoring. RESULTS: The sample panel, validated by National Medical Products Administration (NMPA)-approved tests and targeted bisulfite sequencing, met expectations and could be used for external quality assessment (EQA). Among the 59 laboratories, 55 (93.22%) correctly reported the mSEPT9 results for all samples, while four (6.79%) reported 15 false negatives and were considered improvable. All false negatives originated from four laboratories using laboratory-developed tests (LDTs), with three failing to detect weakly positive samples, samples containing interference, and samples from different CRC cells, and one reported erroneous results on all positive samples. CONCLUSION: Our results illustrated that the detection of mSEPT9 in cfDNA is satisfactory in China. EQA is indispensable because it can help improve the diagnostic capability and quality management of the laboratories, and provide suggestions for the problems existing in mSEPT9 detection.

The cell-end protein Tea4 spatially regulates hyphal branch initiation and appressorium remodeling in the blast fungus Magnaporthe oryzae.

The differentiation of specialized infection cells, called appressoria, from polarized germ tubes of the blast fungus Magnaporthe oryzae, requires remarkable remodeling of cell polarity and architecture, yet our understanding of this process remains incomplete. Here we investigate the behavior and role of cell-end marker proteins in appressorium remodeling and hyphal branch emergence. We show that the SH3 domain-containing protein Tea4 is required for the normal formation of an F-actin ring at Tea1-GFP-labeled polarity nodes, which contributes to the remodeling of septin structures and repolarization of the appressorium. Further, we show that Tea1 localizes to a cortical structure during hyphal septation which, unlike contractile septin rings, persists after septum formation, and, in combination with other polarity determinants, likely spatially regulates branch emergence. Genetic loss of Tea4 leads to mislocalization of Tea1 at the hyphal apex and with it, impaired growth directionality. In contrast, Tea1 is largely depleted from septation events in Δtea4 mutants and branching and septation are significantly reduced. Together, our data provide new insight into polarity remodeling during infection-related and vegetative growth by the blast fungus.

Septins modulate the autophagy response after nutrient starvation.

The pathways that induce macroautophagy (referred to as autophagy hereafter) in response to the stress of starvation are well conserved and essential under nutrient-limiting conditions. However, less is understood about the mechanisms that modulate the autophagy response. Here we present evidence that after induction of autophagy in budding yeast septin filaments rapidly assemble into discrete patches distributed along the cell cortex. These patches gradually mature over 12 h of nutrient deprivation to form extended structures around Atg9 membranes tethered at the cortical endoplasmic reticulum, a class of membranes that are limiting for autophagosome biogenesis. Loss of cortical septin structures alters the kinetics of autophagy activation and most dramatically extends the duration of the autophagy response. In wild-type cells, diffusion of Atg9 membranes at the cell cortex undergoes transient pauses that are dependent on septins, and septins at the bud neck block the diffusion of Atg9 membranes between mother and daughter cells. We conclude that septins reorganize at the cell cortex during autophagy to locally limit access of Atg9 membranes to autophagosome assembly sites, and thus modulate the autophagy response during nutrient deprivation.

The Septin Gene StSep4 Contributes to the Pathogenicity of Setosphaeria turcica by Regulating the Morphology, Cell Wall Integrity, and Pathogenic Factor Biosynthesis.

Septins are a conserved group of GTP-binding proteins found in all eukaryotes and are the fourth-most abundant cytoskeletal proteins. Septins of some pathogenic fungi are involved in morphological changes related to infection. Our previous studies have identified four core septins (StSep1-4) in Setosphaeria turcica, the causal agent of northern corn leaf blight, while only StSep4 is significantly upregulated during the invasive process. We therefore used forchlorfenuron (FCF), the specific inhibitor of septin, and ΔStSep4 knockout mutants to further clarify the role of septins in S. turcica pathogenicity. FCF treatment caused a dose-dependent reduction in S. turcica colony growth, delayed the formation of infection structures, and reduced the penetration ability. ΔStSep4 knockout mutants displayed abnormal mycelium morphology, slow mycelial growth, conidiation deficiency, delayed appressorium development, and weakened pathogenicity. StSep4 deletion also broke cell wall integrity, altered chitin distribution, decreased the melanin content, and disrupted normal nuclear localization. A transcriptomic comparison revealed that genes differentially expressed between ΔStSep4 and WT were enriched in terms of ribosomes, protein translation, membrane components, and transmembrane transport activities. Our results demonstrate that StSep4 is required for morphology and pathogenicity in S. turcica, making it a promising target for the development of novel fungicides.

The value of serum methylated septin 9 and carcinoembryonic antigen in efficacy evaluation and follow-up monitoring of colorectal cancer.

This study explored the value of the detection of serum methylated septin 9 (mSEPT9) and carcinoembryonic antigen (CEA) in the auxiliary diagnosis, curative effect evaluation, and follow-up monitoring of colorectal cancer (CRC). The diagnosis and treatment data of 208 CRC patients in the First Affiliated Hospital of Xinjiang Medical University (China) were collected from March 2019 to December 2019, and these patients were followed up. The correlation between serum CEA, mSEPT9 levels, and tumor location and size were analyzed. Serum mSEPT9 and CEA were detected before and after surgery and during follow-up after treatment to analyze the value of mSEPT9 in efficacy evaluation and follow-up monitoring. In 87 patients with CRC patients who underwent surgery, the average size of poorly differentiated tumors was the largest (25.01±14.08 cm2), which was significantly different from that of moderately differentiated tumors (P =0.039). There was a statistically significant difference in serum CEA level among different degrees of differentiation (P=0.018). The level of CEA was relatively low when tumors occurred in the transverse and ascending colon. When the level of CEA was high, negative mSEPT9 suggested that the probability of a tumor occurring in the cecum was high; positive mSEPT9 indicated that the tumor was highly likely to occur in the descending or sigmoid colon. Detection before and after surgery revealed that the level of mSEPT9 may be related to the tumor-bearing state of patients. A Follow-up study also showed that the sensitivity and specificity of mSEPT9 for recurrence and metastasis were 83.3% and 97.7%, respectively, and the sensitivity and specificity of CEA were 61.1% and 89.5%, respectively. The combined detection of mSEPT9 and CEA can indicate the location and size of colorectal cancer, while the detection of serum mSEPT9 may have clinical significance in the efficacy evaluation and follow-up monitoring of colorectal cancer. KEY WORDS: Colorectal Cancer, mSEPT9, Recurrence, Metastasis, CEA.

Reconstituted in vitro systems to reveal the roles and functions of septins.

Septins are essential cytoskeletal proteins involved in key cellular processes and have also been implicated in diseases from cancers to neurodegenerative pathologies. However, they have not been as thoroughly studied as other cytoskeletal proteins. In vivo, septins interact with other cytoskeletal proteins and with the inner plasma membrane. Hence, bottom-up in vitro cell-free assays are well suited to dissect the roles and behavior of septins in a controlled environment. Specifically, in vitro studies have been invaluable in describing the self-assembly of septins into a large diversity of ultrastructures. Given that septins interact specifically with membrane, the details of these septin-membrane interactions have been analyzed using reconstituted lipid systems. In particular, at a membrane, septins are often localized at curvatures of micrometer scale. In that context, in vitro assays have been performed with substrates of varying curvatures (spheres, cylinders or undulated substrates) to probe the sensitivity of septins to membrane curvature. This Review will first present the structural properties of septins in solution and describe the interplay of septins with cytoskeletal partners. We will then discuss how septins interact with biomimetic membranes and induce their reshaping. Finally, we will highlight the curvature sensitivity of septins and how they alter the mechanical properties of membranes.

Anillin forms linear structures and facilitates furrow ingression after septin and formin depletion.

During cytokinesis, a contractile ring consisting of unbranched filamentous actin (F-actin) and myosin II constricts at the cell equator. Unbranched F-actin is generated by formin, and without formin no cleavage furrow forms. In Caenorhabditis elegans, depletion of septin restores furrow ingression in formin mutants. How the cleavage furrow ingresses without a detectable unbranched F-actin ring is unknown. We report that, in this setting, anillin (ANI-1) forms a meshwork of circumferentially aligned linear structures decorated by non-muscle myosin II (NMY-2). Analysis of ANI-1 deletion mutants reveals that its disordered N-terminal half is required for linear structure formation and sufficient for furrow ingression. NMY-2 promotes the circumferential alignment of the linear ANI-1 structures and interacts with various lipids, suggesting that NMY-2 links the ANI-1 network with the plasma membrane. Collectively, our data reveal a compensatory mechanism, mediated by ANI-1 linear structures and membrane-bound NMY-2, that promotes furrowing when unbranched F-actin polymerization is compromised.

Regulation of integrin α5ß1-mediated Staphylococcus aureus cellular invasion by the septin cytoskeleton.

Staphylococcus aureus, a Gram-positive bacterial pathogen, is an urgent health threat causing a wide range of clinical infections. Originally viewed as a strict extracellular pathogen, accumulating evidence has revealed S. aureus to be a facultative intracellular pathogen subverting host cell signalling to support invasion. The majority of clinical isolates produce fibronectin-binding proteins A and B (FnBPA and FnBPB) to interact with host integrin α5ß1, a key component of focal adhesions. S. aureus binding of integrin α5ß1 promotes its clustering on the host cell surface, triggering activation of focal adhesion kinase (FAK) and cytoskeleton rearrangements to promote bacterial invasion into non-phagocytic cells. Here, we discover that septins, a component of the cytoskeleton that assembles on membranes, are recruited as collar-like structures with actin to S. aureus invasion sites engaging integrin α5ß1. To investigate septin recruitment to the plasma membrane in a bacteria-free system, we used FnBPA-coated latex beads and showed that septins are recruited upon activation of integrin α5ß1. SEPT2 depletion reduced S. aureus invasion, but increased surface expression of integrin α5 and adhesion of S. aureus to host cells. Consistent with this, SEPT2 depletion increased cellular protein levels of integrin α5 and ß1 subunits, as well as FAK. Collectively, these results provide insights into regulation of integrin α5ß1 and invasion of S. aureus by the septin cytoskeleton.

Mechanistic model for nuclear migration in hyphae during mitosis.

Saccharomyces cerevisiae and Candida albicans, the two well-known human pathogens, can be found in all three morphologies, i.e., yeast, pseudohyphae, and true hyphae. The cylindrical daughter-bud (germ tube) grows very long for true hyphae, and the cell cycle is delayed compared to the other two morphologies. The place of the nuclear division is specific for true hyphae determined by the position of the septin ring. However, the septin ring can localize anywhere inside the germ tube, unlike the mother-bud junction in budding yeast. Since the nucleus often migrates a long path in the hyphae, the underlying mechanism must be robust for executing mitosis in a timely manner. We explore the mechanism of nuclear migration through hyphae in light of mechanical interactions between astral microtubules and the cell cortex. We report that proper migration through constricted hyphae requires a large dynein pull applied on the astral microtubules from the hyphal cortex. This is achieved when the microtubules frequently slide along the hyphal cortex so that a large population of dyneins actively participate, pulling on them. Simulation shows timely migration when the dyneins from the mother cortex do not participate in pulling on the microtubules. These findings are robust for long migration and positioning of the nucleus in the germ tube at the septin ring.

Nanoscale details of mitochondrial constriction revealed by cryoelectron tomography.

Mitochondria adapt to changing cellular environments, stress stimuli, and metabolic demands through dramatic morphological remodeling of their shape, and thus function. Such mitochondrial dynamics is often dependent on cytoskeletal filament interactions. However, the precise organization of these filamentous assemblies remains speculative. Here, we apply cryogenic electron tomography to directly image the nanoscale architecture of the cytoskeletal-membrane interactions involved in mitochondrial dynamics in response to damage. We induced mitochondrial damage via membrane depolarization, a cellular stress associated with mitochondrial fragmentation and mitophagy. We find that, in response to acute membrane depolarization, mammalian mitochondria predominantly organize into tubular morphology that abundantly displays constrictions. We observe long bundles of both unbranched actin and septin filaments enriched at these constrictions. We also observed septin-microtubule interactions at these sites and elsewhere, suggesting that these two filaments guide each other in the cytosolic space. Together, our results provide empirical parameters for the architecture of mitochondrial constriction factors to validate/refine existing models and inform the development of new ones.

Methylated Septin9 identified patients with colorectal carcinoma and showed higher sensitivity than conventional biomarkers in detecting tumor.

INTRODUCTION: It is worth noting the limitations in sensitivity of the existing biomarkers carcinoembryonic antigen (CEA) and carbohydrate antigen (CA 19-9) in detection of colorectal cancer (CRC). In our study, we address the performance of the liquid biopsy biomarker "methylated septin 9" (mSEPT9) in the detection and disease surveillance of CRC. MATERIALS AND METHODS: The monocentric prospective survey encompassed 120 patients diagnosed with CRC who underwent planned curative resection between December 2018 and December 2020. Blood samples were collected from the participants preoperatively as well as at 7 days, 6 weeks, and 3 months postoperatively. The presence of mSEPT9, CEA, and CA 19-9 was detected using the pro Epi Colon® 2.0 CE test, Elecsys® CEA, and Elecsys® CA19-9 electrochemiluminescence immunoassay, respectively. RESULTS: In the preoperative setting, mSEPT9 demonstrated superior capability in identifying patients with CRC compared to CEA and CA 19-9, with detection rates of 57%, 32%, and 18% respectively. Combining all three biomarkers increased the overall sensitivity to 66% preoperatively. In considering UICC stage and T-status, mSEPT9 exhibited higher sensitivity across all stages in comparison with conventional tumor markers, and 65% of patients with metastases were identified postoperatively through mSEPT9. Tumor recognition after surgery was achieved with the sensitivity of 72% and specificity of 91%. CONCLUSIONS: We recommend using mSEPT9 as a non-invasive diagnostic tool for the ongoing monitoring of patients with CRC. The sensitivity and specificity exhibited by mSEPT9 in recognition of tumor after surgery, highlights its particular potential for monitoring of CRC patients.

Macrophage-to-endothelial cell crosstalk by the cholesterol metabolite 27HC promotes atherosclerosis in male mice.

Hypercholesterolemia and vascular inflammation are key interconnected contributors to the pathogenesis of atherosclerosis. How hypercholesterolemia initiates vascular inflammation is poorly understood. Here we show in male mice that hypercholesterolemia-driven endothelial activation, monocyte recruitment and atherosclerotic lesion formation are promoted by a crosstalk between macrophages and endothelial cells mediated by the cholesterol metabolite 27-hydroxycholesterol (27HC). The pro-atherogenic actions of macrophage-derived 27HC require endothelial estrogen receptor alpha (ERα) and disassociation of the cytoplasmic scaffolding protein septin 11 from ERα, leading to extranuclear ERα- and septin 11-dependent activation of NF-κB. Furthermore, pharmacologic inhibition of cyp27a1, which generates 27HC, affords atheroprotection by reducing endothelial activation and monocyte recruitment. These findings demonstrate cell-to-cell communication by 27HC, and identify a major causal linkage between the hypercholesterolemia and vascular inflammation that partner to promote atherosclerosis. Interventions interrupting this linkage may provide the means to blunt vascular inflammation without impairing host defense to combat the risk of atherosclerotic cardiovascular disease that remains despite lipid-lowering therapies.

Cyclophilin A Isomerisation of Septin 2 Mediates Abscission during Cytokinesis.

The isomerase activity of Cyclophilin A is important for midbody abscission during cell division, however, to date, midbody substrates remain unknown. In this study, we report that the GTP-binding protein Septin 2 interacts with Cyclophilin A. We highlight a dynamic series of Septin 2 phenotypes at the midbody, previously undescribed in human cells. Furthermore, Cyclophilin A depletion or loss of isomerase activity is sufficient to induce phenotypic Septin 2 defects at the midbody. Structural and molecular analysis reveals that Septin 2 proline 259 is important for interaction with Cyclophilin A. Moreover, an isomerisation-deficient EGFP-Septin 2 proline 259 mutant displays defective midbody localisation and undergoes impaired abscission, which is consistent with data from cells with loss of Cyclophilin A expression or activity. Collectively, these data reveal Septin 2 as a novel interacting partner and isomerase substrate of Cyclophilin A at the midbody that is required for abscission during cytokinesis in cancer cells.