Pacsin 2-dependent N-cadherin internalization regulates the migration behaviour of malignant cancer cells.

Collective cell migration is the coordinated movement of multiple cells connected by cadherin-based adherens junctions and is essential for physiological and pathological processes. Cadherins undergo dynamic intracellular trafficking, and their surface level is determined by a balance between endocytosis, recycling and degradation. However, the regulatory mechanism of cadherin turnover in collective cell migration remains elusive. In this study, we show that the Bin/amphiphysin/Rvs (BAR) domain protein pacsin 2 (protein kinase C and casein kinase substrate in neurons protein 2) plays an essential role in collective cell migration by regulating N-cadherin (also known as CDH2) endocytosis in human cancer cells. Pacsin 2-depleted cells formed cell-cell contacts enriched with N-cadherin and migrated in a directed manner. Furthermore, pacsin 2-depleted cells showed attenuated internalization of N-cadherin from the cell surface. Interestingly, GST pull-down assays demonstrated that the pacsin 2 SH3 domain binds to the cytoplasmic region of N-cadherin, and expression of an N-cadherin mutant defective in binding to pacsin 2 phenocopied pacsin 2 RNAi cells both in cell contact formation and N-cadherin endocytosis. These data support new insights into a novel endocytic route of N-cadherin in collective cell migration, highlighting pacsin 2 as a possible therapeutic target for cancer metastasis.

Tip-scan high-speed atomic force microscopy with a uniaxial substrate stretching device for studying dynamics of biomolecules under mechanical stress.

High-speed atomic force microscopy (HS-AFM) is a powerful tool for studying the dynamics of biomolecules in vitro because of its high temporal and spatial resolution. However, multi-functionalization, such as combination with complementary measurement methods, environment control, and large-scale mechanical manipulation of samples, is still a complex endeavor due to the inherent design and the compact sample scanning stage. Emerging tip-scan HS-AFM overcame this design hindrance and opened a door for additional functionalities. In this study, we designed a motor-driven stretching device to manipulate elastic substrates for HS-AFM imaging of biomolecules under controllable mechanical stimulation. To demonstrate the applicability of the substrate stretching device, we observed a microtubule buckling by straining the substrate and actin filaments linked by α-actinin on a curved surface. In addition, a BAR domain protein BIN1 that senses substrate curvature was observed while dynamically controlling the surface curvature. Our results clearly prove that large-scale mechanical manipulation can be coupled with nanometer-scale imaging to observe biophysical effects otherwise obscured.

Centronuclear Myopathy Caused by Defective Membrane Remodelling of Dynamin 2 and BIN1 Variants.

Centronuclear myopathy (CNM) is a congenital myopathy characterised by centralised nuclei in skeletal myofibers. T-tubules, sarcolemmal invaginations required for excitation-contraction coupling, are disorganised in the skeletal muscles of CNM patients. Previous studies showed that various endocytic proteins are involved in T-tubule biogenesis and their dysfunction is tightly associated with CNM pathogenesis. DNM2 and BIN1 are two causative genes for CNM that encode essential membrane remodelling proteins in endocytosis, dynamin 2 and BIN1, respectively. In this review, we overview the functions of dynamin 2 and BIN1 in T-tubule biogenesis and discuss how their dysfunction in membrane remodelling leads to CNM pathogenesis.

The Lipid-Binding Defective Dynamin 2 Mutant in Charcot-Marie-Tooth Disease Impairs Proper Actin Bundling and Actin Organization in Glomerular Podocytes.

Dynamin is an endocytic protein that functions in vesicle formation by scission of invaginated membranes. Dynamin maintains the structure of foot processes in glomerular podocytes by directly and indirectly interacting with actin filaments. However, molecular mechanisms underlying dynamin-mediated actin regulation are largely unknown. Here, biochemical and cell biological experiments were conducted to uncover how dynamin modulates interactions between membranes and actin in human podocytes. Actin-bundling, membrane tubulating, and GTPase activities of dynamin were examined in vitro using recombinant dynamin 2-wild-type (WT) or dynamin 2-K562E, which is a mutant found in Charcot-Marie-Tooth patients. Dynamin 2-WT and dynamin 2-K562E led to the formation of prominent actin bundles with constant diameters. Whereas liposomes incubated with dynamin 2-WT resulted in tubule formation, dynamin 2-K562E reduced tubulation. Actin filaments and liposomes stimulated dynamin 2-WT GTPase activity by 6- and 20-fold, respectively. Actin-filaments, but not liposomes, stimulated dynamin 2-K562E GTPase activity by 4-fold. Self-assembly-dependent GTPase activity of dynamin 2-K562E was reduced to one-third compared to that of dynamin 2-WT. Incubation of liposomes and actin with dynamin 2-WT led to the formation of thick actin bundles, which often bound to liposomes. The interaction between lipid membranes and actin bundles by dynamin 2-K562E was lower than that by dynamin 2-WT. Dynamin 2-WT partially colocalized with stress fibers and actin bundles based on double immunofluorescence of human podocytes. Dynamin 2-K562E expression resulted in decreased stress fiber density and the formation of aberrant actin clusters. Dynamin 2-K562E colocalized with α-actinin-4 in aberrant actin clusters. Reformation of stress fibers after cytochalasin D-induced actin depolymerization and washout was less effective in dynamin 2-K562E-expressing cells than that in dynamin 2-WT. Bis-T-23, a dynamin self-assembly enhancer, was unable to rescue the decreased focal adhesion numbers and reduced stress fiber density induced by dynamin 2-K562E expression. These results suggest that the low affinity of the K562E mutant for lipid membranes, and atypical self-assembling properties, lead to actin disorganization in HPCs. Moreover, lipid-binding and self-assembly of dynamin 2 along actin filaments are required for podocyte morphology and functions. Finally, dynamin 2-mediated interactions between actin and membranes are critical for actin bundle formation in HPCs.

Imaging-based evaluation of pathogenicity by novel DNM2 variants associated with centronuclear myopathy.

A centronuclear myopathy (CNM) is a group of inherited congenital diseases showing clinically progressive muscle weakness associated with the presence of centralized myonuclei, diagnosed by genetic testing and muscle biopsy. The gene encoding dynamin 2, DNM2, has been identified as a causative gene for an autosomal dominant form of CNM. However, the information of a DNM2 variant alone is not always sufficient to gain a definitive diagnosis as the pathogenicity of many gene variants is currently unknown. In this study, we identified five novel DNM2 variants in our cohort. To establish the pathogenicity of these variants without using clinicopathological information, we used a simple in cellulo imaging-based assay for T-tubule-like structures to provide quantitative data that enable objective determination of pathogenicity by novel DNM2 variants. With this assay, we demonstrated that the phenotypes induced by mutant dynamin 2 in cellulo are well correlated with biochemical gain-of-function features of mutant dynamin 2 as well as the clinicopathological phenotypes of each patient. Our approach of combining an in cellulo assay with clinical information of the patients also explains the course of a disease progression by the pathogenesis of each variant in DNM2-associated CNM.

Dynamin 2 and BAR domain protein pacsin 2 cooperatively regulate formation and maturation of podosomes.

Podosomes are actin-rich adhesion structures formed in a variety of cell types, such as monocytic cells or cancer cells, to facilitate attachment to and degradation of the extracellular matrix (ECM). Previous studies showed that dynamin 2, a large GTPase involved in membrane remodeling and actin organization, is required for podosome function. However, precise roles of dynamin 2 at the podosomes remain to be elucidated. In this study, we identified a BAR (Bin-Amphiphysin-Rvs167) domain protein pacsin 2 as a functional partner of dynamin 2 at podosomes. Dynamin 2 and pacsin 2 interact and co-localize to podosomes in Src-transformed NIH 3T3 (NIH-Src) cells. RNAi of either dynamin 2 or pacsin 2 in NIH-Src cells inhibited podosome formation and maturation, suggesting essential and related roles at podosomes. Consistently, RNAi of pacsin 2 prevented dynamin 2 localization to podosomes, and reciprocal RNAi of dynamin 2 prevented pacsin 2 localization to podosomes. Taking these results together, we conclude that dynamin 2 and pacsin 2 co-operatively regulate organization of podosomes in NIH-Src cells.

Nucleation process of the 2011 northern Nagano earthquake from nearby seismic observations.

The 2011 magnitude (M) 9.0 Tohoku-oki earthquake was followed by seismicity activation in inland areas throughout Japan. An outstanding case is the M6.2 Northern Nagano earthquake, central Japan, occurred 13-h after the megathrust event, approximately 400 km away from its epicenter. The physical processes relating the occurrence of megathrust earthquakes and subsequent activation of relatively large inland earthquakes are not well understood. Here we use waveform data of a dense local seismic network to reveal with an unprecedented resolution the complex mechanisms leading to the occurrence of the M6.2 earthquake. We show that previously undetected small earthquakes initiated along the Nagano earthquake source fault at relatively short times after the Tohoku-oki megathrust earthquake, and the local seismicity continued intermittently until the occurrence of the M6.2 event, being likely 'modulated' by the arrival of surface waves from large, remote aftershocks off-shore Tohoku. About 1-h before the Nagano earthquake, there was an acceleration of micro-seismicity migrating towards its hypocenter. Migration speeds indicate potential localized slow-slip, culminating with the occurrence of the large inland earthquake, with fluids playing a seismicity-activation role at a regional scale.

Mutant BIN1-Dynamin 2 complexes dysregulate membrane remodeling in the pathogenesis of centronuclear myopathy.

Membrane remodeling is required for dynamic cellular processes such as cell division, polarization, and motility. BAR domain proteins and dynamins are key molecules in membrane remodeling that work together for membrane deformation and fission. In striated muscles, sarcolemmal invaginations termed T-tubules are required for excitation-contraction coupling. BIN1 and DNM2, which encode a BAR domain protein BIN1 and dynamin 2, respectively, have been reported to be causative genes of centronuclear myopathy (CNM), a hereditary degenerative disease of skeletal muscle, and deformation of T-tubules is often observed in the CNM patients. However, it remains unclear how BIN1 and dynamin 2 are implicated in T-tubule biogenesis and how mutations in these molecules cause CNM to develop. Here, using an in cellulo reconstitution assay, we demonstrate that dynamin 2 is required for stabilization of membranous structures equivalent to T-tubules. GTPase activity of wild-type dynamin 2 is suppressed through interaction with BIN1, whereas that of the disease-associated mutant dynamin 2 remains active due to lack of the BIN1-mediated regulation, thus causing aberrant membrane remodeling. Finally, we show that in cellulo aberrant membrane remodeling by mutant dynamin 2 variants is correlated with their enhanced membrane fission activities, and the results can explain severity of the symptoms in patients. Thus, this study provides molecular insights into dysregulated membrane remodeling triggering the pathogenesis of DNM2-related CNM.

Dynamin 1 is important for microtubule organization and stabilization in glomerular podocytes.

Dynamin 1 is a neuronal endocytic protein that participates in vesicle formation by scission of invaginated membranes. Dynamin 1 is also expressed in the kidney; however, its physiological significance to this organ remains unknown. Here, we show that dynamin 1 is crucial for microtubule organization and stabilization in glomerular podocytes. By immunofluorescence and immunoelectron microscopy, dynamin 1 was concentrated at microtubules at primary processes in rat podocytes. By immunofluorescence of differentiated mouse podocytes (MPCs), dynamin 1 was often colocalized with microtubule bundles, which radially arranged toward periphery of expanded podocyte. In dynamin 1-depleted MPCs by RNAi, α-tubulin showed a dispersed linear filament-like localization, and microtubule bundles were rarely observed. Furthermore, dynamin 1 depletion resulted in the formation of discontinuous, short acetylated α-tubulin fragments, and the decrease of microtubule-rich protrusions. Dynamins 1 and 2 double-knockout podocytes showed dispersed acetylated α-tubulin and rare protrusions. In vitro, dynamin 1 polymerized around microtubules and cross-linked them into bundles, and increased their resistance to the disassembly-inducing reagents Ca2+ and podophyllotoxin. In addition, overexpression and depletion of dynamin 1 in MPCs increased and decreased the nocodazole resistance of microtubules, respectively. These results suggest that dynamin 1 supports the microtubule bundle formation and participates in the stabilization of microtubules.

[Corrigendum]Phosphorylation of cortactin by cyclin-dependent kinase 5 modulates actin bundling by the dynamin 1-cortactin ring-like complex and formation of filopodia and lamellipodia in NG108-15 glioma-derived cells.

Subsequently to the publication of the above article, the authors have realized that the second­listed author, The Mon La, had not been properly credited as one of the co­writers of the paper. Therefore, the Authors' Contributions of the Declarations section of the article should have read as follows: Authors' contributions HY, KTa and TML designed the research and wrote the paper. HY, TA, YM, EO and TT performed mutant protein construction, protein purification and actin bundling experiments. TA and YM performed electron microscopy. EO, TML, KS and KF performed immunofluorescent microscopy, cell migration assay and analyzed data. FYW and KTo identified phosphorylation sites by MALDI­MS. All authors read and approved the final manuscript. The authors apologize to the readership of the Journal for the misinformation in this regard, and for any inconvenience caused. [the original article was published in International Journal of Oncology 54: 550­558, 2019; DOI: 10.3892/ijo.2018.4663].

Phosphorylation of cortactin by cyclin-dependent kinase 5 modulates actin bundling by the dynamin 1-cortactin ring-like complex and formation of filopodia and lamellipodia in NG108-15 glioma-derived cells.

Dynamin copolymerizes with cortactin to form a ring­like complex that bundles and stabilizes actin filaments. Actin bundle formation is crucial for generation of filopodia and lamellipodia, which guide migration, invasion, and metastasis of cancer cells. However, it is unknown how the dynamin­cortactin complex regulates actin bundle formation. The present study investigated phosphorylation of cortactin by cyclin­dependent kinase 5 (CDK5) and its effect on actin bundle formation by the dynamin­cortactin complex. CDK5 directly phosphorylated cortactin at T145/T219 in vitro. Phosphomimetic mutants in which one or both of these threonine residues was substituted by aspartate were used. The three phosphomimetic mutants (T145D, T219D and T145DT219D) had a decreased affinity for F­actin. Furthermore, electron microscopy demonstrated that these phosphomimetic mutants could not form a ring­like complex with dynamin 1. Consistently, the dynamin 1­phosphomimetic cortactin complexes exhibited decreased actin­bundling activity. Expression of the phosphomimetic mutants resulted in not only aberrant lamellipodia and short filopodia but also cell migration in NG108­15 glioma­derived cells. These results indicate that phosphorylation of cortactin by CDK5 regulates formation of lamellipodia and filopodia by modulating dynamin 1/cortactin­dependent actin bundling. Taken together, these findings suggest that CDK5 is a potential molecular target for anticancer therapy.

Dynamic clustering of dynamin-amphiphysin helices regulates membrane constriction and fission coupled with GTP hydrolysis.

Dynamin is a mechanochemical GTPase essential for membrane fission during clathrin-mediated endocytosis. Dynamin forms helical complexes at the neck of clathrin-coated pits and their structural changes coupled with GTP hydrolysis drive membrane fission. Dynamin and its binding protein amphiphysin cooperatively regulate membrane remodeling during the fission, but its precise mechanism remains elusive. In this study, we analyzed structural changes of dynamin-amphiphysin complexes during the membrane fission using electron microscopy (EM) and high-speed atomic force microscopy (HS-AFM). Interestingly, HS-AFM analyses show that the dynamin-amphiphysin helices are rearranged to form clusters upon GTP hydrolysis and membrane constriction occurs at protein-uncoated regions flanking the clusters. We also show a novel function of amphiphysin in size control of the clusters to enhance biogenesis of endocytic vesicles. Our approaches using combination of EM and HS-AFM clearly demonstrate new mechanistic insights into the dynamics of dynamin-amphiphysin complexes during membrane fission.

Down-dip variations in a subducting low-velocity zone linked to episodic tremor and slip: a new constraint from ScSp waves.

Fluids are thought to play an important role in controlling episodic tremor and slow slip (ETS) in subduction zones. Therefore, constraining the along-dip distribution of fluids is necessary to better understand source mechanism of ETS, and particularly the role played by fluids in ETS generation. Here, we report clear observations of coherent ScSp phases with a dense seismic array in western Shikoku, Japan, where ETS has been most active over the past decade. Using numerical simulations of elastic-wave propagation to reproduce the observed ScSp phases, we demonstrate that, relative to shallower depths, either the Vp/Vs ratio or the thickness of a low-velocity zone (LVZ) within the subducting oceanic crust increases with depth beneath the mantle wedge corner where ETS has been observed. Based on these depth dependences of the structural elements, a wide semi-ductile shear zone appears to be lubricated by high-pressurized fluid in the subducting oceanic crust at ETS source depths, and to be a key factor regulating ETS activity.

Organophosphate flame retardants in the indoor air and dust in cars in Japan.

The concentrations of organophosphate flame retardants (OPFRs) in the indoor air and dust were measured in 25 unoccupied cars in Japan. In the indoor air of the cars, most OPFRs were neither detected nor found at a concentration lower than the method quantification limit. The highest concentration (1500 ng m-3) was obtained for tris(1-chloro-2-propyl) phosphate (TCIPP). By contrast, many OPFRs were detected in the dust samples collected from the interior of the cars. TCIPP and tris(2-ethylhexyl) phosphate (TEHP) were present at the highest concentrations at 390 µg g-1 (in dust from car seats) and 640 µg g-1 (in dust from car floor mats), respectively. The highest median concentrations (35 µg g-1 for car seats, 53 µg g-1 for car floor mats) were obtained for tris(2-butoxyethyl) phosphate (TBOEP). According to the results of our exposure assessment, the typical exposures to OPFRs via inhalation in car cabins ranged from 9.0×10-4 to 7.8×10-1 ng kg-bw-1 day-1. The typical exposures to OPFRs via dust ingestion ranged from 9.2×10-4 to 8.8×10-1 ng kg-bw-1 day-1. We compared these results with the ref-erence doses for OPFRs and found that, based on cur-rent information about the toxicities of OPFRs, exposure to OPFRs in car cabins via inhalation and dust ingestion is unlikely to have adverse human health effects.

Dynamin2 GTPase contributes to invadopodia formation in invasive bladder cancer cells.

Cancer cell invasion is mediated by actin-based membrane protrusions termed invadopodia. Invadopodia consist of "core" F-actin bundles associated with adhesive and proteolytic machineries promoting cell invasion by degrading extracellular matrix (ECM). Formation of the F-actin core in invadopodia is regulated by various actin-binding proteins including Arp2/3 complex and cortactin. Dynamin GTPase localizes to the invadopodia and is implicated in cancer cell invasion, but its precise role at the invadopodia remained elusive. In this study, we examined the roles of dynamin at the invadopodia of bladder cancer cells. Although all three dynamin isoforms (dynamin1, 2 and 3) are expressed in human bladder cancer cell line T24, only dynamin2 localizes to the invadopodia. Inhibition of dynamin2 function, using either RNA interference (RNAi) or the dynamin specific inhibitor Dynasore, caused defects in invadopodia formation and suppressed invasive activity of T24 bladder cancer cells. Structure-function analysis using dynamin2 deletion fragments identified the proline/arginine-rich domain (PRD) of dynamin2 as indispensable for invadopodia formation and invasiveness of T24 cells. Thus, dynamin2 contributes to bladder cancer invasion by controlling invadopodia formation in bladder cancer cells and may prove a valuable therapeutic target.

Actin bundling by dynamin 2 and cortactin is implicated in cell migration by stabilizing filopodia in human non-small cell lung carcinoma cells.

The endocytic protein dynamin participates in the formation of actin-based membrane protrusions such as podosomes, pseudopodia, and invadopodia, which facilitate cancer cell migration, invasion, and metastasis. However, the role of dynamin in the formation of actin-based membrane protrusions at the leading edge of cancer cells is unclear. In this study, we demonstrate that the ubiquitously expressed dynamin 2 isoform facilitates cell migration by stabilizing F-actin bundles in filopodia of the lung cancer cell line H1299. Pharmacological inhibition of dynamin 2 decreased cell migration and filopodial formation. Furthermore, dynamin 2 and cortactin mostly colocalized along F-actin bundles in filopodia of serum-stimulated H1299 cells by immunofluorescent and immunoelectron microscopy. Knockdown of dynamin 2 or cortactin inhibited the formation of filopodia in serum-stimulated H1299 cells, concomitant with a loss of F-actin bundles. Expression of wild-type cortactin rescued the punctate-like localization of dynamin 2 and filopodial formation. The incubation of dynamin 2 and cortactin with F-actin induced the formation of long and thick actin bundles, with these proteins colocalizing at F-actin bundles. A depolymerization assay revealed that dynamin 2 and cortactin increased the stability of F-actin bundles. These results indicate that dynamin 2 and cortactin participate in cell migration by stabilizing F-actin bundles in filopodia. Taken together, these findings suggest that dynamin might be a possible molecular target for anticancer therapy.

Expression of a dynamin 2 mutant associated with Charcot-Marie-Tooth disease leads to aberrant actin dynamics and lamellipodia formation.

Specific mutations in dynamin 2 are linked to Charcot-Marie-Tooth disease (CMT), an inherited peripheral neuropathy. However, the effects of these mutations on dynamin function, particularly in relation to the regulation of the actin cytoskeleton remain unclear. Here, selected CMT-associated dynamin mutants were expressed to examine their role in the pathogenesis of CMT in U2OS cells. Ectopic expression of the dynamin CMT mutants 555Δ3 and K562E caused an approximately 50% decrease in serum stimulation-dependent lamellipodia formation; however, only K562E caused aberrations in the actin cytoskeleton. Immunofluorescence analysis showed that the K562E mutation resulted in the disappearance of radially aligned actin bundles and the simultaneous appearance of F-actin clusters. Live-cell imaging analyses showed F-actin polymers of decreased length assembled into immobile clusters in K562E-expressing cells. The K562E dynamin mutant colocalized with the F-actin clusters, whereas its colocalization with clathrin-coated pit marker proteins was decreased. Essentially the same results were obtained using another cell line, HeLa and NG108-15 cells. The present study is the first to show the association of dynamin CMT mutations with aberrant actin dynamics and lamellipodia, which may contribute to defective endocytosis and myelination in Schwann cells in CMT.

Possible role of cortactin phosphorylation by protein kinase Cα in actin-bundle formation at growth cone.

BACKGROUND INFORMATION: Cortactin contributes to growth cone morphogenesis by forming with dynamin, ring-shaped complexes that mechanically bundle and stabilise F-actin. However, the regulatory mechanism of cortactin action is poorly understood. RESULTS: Immunofluorescence microscopy revealed that protein kinase C (PKC) α colocalises with cortactin at growth cone filopodia in SH-SY5Y neuroblastoma cells. PKC activation by phorbol 12-myristate 13-acetate causes cortactin phosphorylation, filopodial retraction and F-actin-bundle loss. Moreover, PKCα directly phosphorylates cortactin in vitro at S135/T145/S172, mitigating both cortactin's actin-binding and actin-crosslinking activity, whereas cellular expression of a phosphorylation-mimetic cortactin mutant hinders filopodial formation with a significant decrease of actin bundles. CONCLUSIONS: Our results indicate that PKC-mediated cortactin phosphorylation might be implicated in the maintenance of growth cone.

Drosophila F-BAR protein Syndapin contributes to coupling the plasma membrane and contractile ring in cytokinesis.

Cytokinesis is a highly ordered cellular process driven by interactions between central spindle microtubules and the actomyosin contractile ring linked to the dynamic remodelling of the plasma membrane. The mechanisms responsible for reorganizing the plasma membrane at the cell equator and its coupling to the contractile ring in cytokinesis are poorly understood. We report here that Syndapin, a protein containing an F-BAR domain required for membrane curvature, contributes to the remodelling of the plasma membrane around the contractile ring for cytokinesis. Syndapin colocalizes with phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) at the cleavage furrow, where it directly interacts with a contractile ring component, Anillin. Accordingly, Anillin is mislocalized during cytokinesis in Syndapin mutants. Elevated or diminished expression of Syndapin leads to cytokinesis defects with abnormal cortical dynamics. The minimal segment of Syndapin, which is able to localize to the cleavage furrow and induce cytokinesis defects, is the F-BAR domain and its immediate C-terminal sequences. Phosphorylation of this region prevents this functional interaction, resulting in reduced ability of Syndapin to bind to and deform membranes. Thus, the dephosphorylated form of Syndapin mediates both remodelling of the plasma membrane and its proper coupling to the cytokinetic machinery.

The chromosomal passenger complex controls the function of endosomal sorting complex required for transport-III Snf7 proteins during cytokinesis.

Cytokinesis controls the proper segregation of nuclear and cytoplasmic materials at the end of cell division. The chromosomal passenger complex (CPC) has been proposed to monitor the final separation of the two daughter cells at the end of cytokinesis in order to prevent cell abscission in the presence of DNA at the cleavage site, but the precise molecular basis for this is unclear. Recent studies indicate that abscission could be mediated by the assembly of filaments comprising components of the endosomal sorting complex required for transport-III (ESCRT-III). Here, we show that the CPC subunit Borealin interacts directly with the Snf7 components of ESCRT-III in both Drosophila and human cells. Moreover, we find that the CPC's catalytic subunit, Aurora B kinase, phosphorylates one of the three human Snf7 paralogues-CHMP4C-in its C-terminal tail, a region known to regulate its ability to form polymers and associate with membranes. Phosphorylation at these sites appears essential for CHMP4C function because their mutation leads to cytokinesis defects. We propose that CPC controls abscission timing through inhibition of ESCRT-III Snf7 polymerization and membrane association using two concurrent mechanisms: interaction of its Borealin component with Snf7 proteins and phosphorylation of CHMP4C by Aurora B.