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Diseases caused byLeishmania, andTrypanosomaparasites, such as leishmaniasis and African sleeping sickness, are a major health problem in tropical countries. Due to their complex life cycle involving both vertebrate and insect hosts, and > 1 billion years of evolutionarily distance, the cell biology of these trypanosomatid parasites exhibits pronounced differences to animal cells. For example, the actin cytoskeleton of trypanosomatids is highly divergent when compared to the other eukaryotes. To understand how actin dynamics are regulated in trypanosomatid parasites, we focused on a central actin-binding protein profilin. Co-crystal structure ofLeishmania majoractin in complex withL. majorprofilin revealed that, although the overall folds of actin and profilin are conserved in eukaryotes,Leishmaniaprofilin contains a unique α-helical insertion, which interacts with the target binding cleft of actin monomer. This insertion is conserved across the Trypanosomatidae family, and is strikingly similar to the structure of WH2 domain, a small actin-binding motif found in many other cytoskeletal regulators. We demonstrate that the WH2-like motif contributes to actin monomer-binding and enhances the actin nucleotide exchange activity ofLeishmaniaprofilin. Surprisingly, unlike other profilins characterized so far,Leishmaniaprofilin inhibited formin-catalyzed actin filament assembly in a mechanism that is dependent on the presence of the WH2-like motif. By generating profilin knockout and knockinLeishmania mexicanastrains, we show that profilin is important for efficient endocytic sorting in parasites, and that the ability to bind actin monomers and proline-rich proteins, as well as the presence of a functional WH2-like motif, are important for thein vivofunction ofLeishmaniaprofilin. Collectively, this study uncovers the molecular principles by which actin dynamics are regulated by profilin in trypanosomatids. Moreover, the unusual actin-binding mechanism of profilin identified here could be applied for designing inhibitors against pathogenic trypanosomatid parasites.AUTHOR SUMMARYLeishmaniaandTrypanosomaparasites are a major health problem as they cause various diseases in humans and other vertebrates. Currently, there are no specific drugs to treat the diseases caused by these trypanosomatid parasites. Similar to all other eukaryotes, trypanosomatid parasites have an actin cytoskeleton, which is essential for the viability of parasites. Interestingly, both actin and actin-regulatory machineries of these parasites are highly divergent from the ones of animals, making them possible drug targets to treat diseases caused by these parasites. To uncover how the actin cytoskeleton of trypanosomatid parasites is regulated, we focused on a central actin-binding protein, profilin. Importantly, our experiments revealed that trypanosomatid profilins interact with actin through a different structural mechanism as compared to animal profilins, and have specific effects on the assembly of actin filaments. Our genetic studies demonstrate that these specific features of trypanosomatid profilin are also critical for the proper function on this protein in parasites. Our study provides new insight into the cell biology of trypanosomatid parasites. We also envision that the structural and functional differences between trypanosomatid and human profilins can be applied for developing compounds for selectively neutralizingLeishmaniaandTrypanosomaparasites.

SUMMARYThe actin cytoskeleton is critical for cell migration, morphogenesis, endocytosis, organelle dynamics, and cytokinesis. To support diverse cellular processes, actin filaments form a variety of structures with specific architectures and dynamic properties. Key proteins specifying actin filaments are tropomyosins. Non-muscle cells express several functionally non-redundant tropomyosin isoforms, which differentially control the interactions of other proteins, including myosins and ADF/cofilin, with actin filaments. However, the underlying molecular mechanisms have remained elusive. By determining the cryogenic electron microscopy structures of actin filaments decorated by two functionally distinct non-muscle tropomyosin isoforms, Tpm1.6 and Tpm3.2, we reveal that actin filament conformation remains unaffected upon binding. However, Tpm1.6 and Tpm3.2 follow different paths along the major groove of the actin filament, providing an explanation for their incapability to co-polymerize on actin filaments. The structures and biochemical work also elucidate the molecular basis underlying specific roles of Tpm1.6 and Tpm3.2 in myosin II activation and protecting actin filaments from ADF/cofilin-catalysed severing.

Funding Information: Acknowledgments: The computations were supported by the University of Chicago Research Funding Information: The computations were supported by the University of Chicago Research Computing Center (RCC). We thank E. Coudrier and C. Simon for insightful discussions. We also thank F. Di Federico for handling plasmids, F. Tabarin-Cayrac for cell sorting, and A.-S. Mace for ImageJ programming assistance. F.-C.T., C.L.C., and P.B. are members of the CNRS consortium AQV. F.-C.T. and P.B. are members of the Labex Cell(n)Scale (ANR-11-LABX0038) and Paris Sciences et Lettres (ANR-10-IDEX-0001-02). We acknowledge the Cell and Tissue Imaging Core facility (PICT IBiSA), Institut Curie, member of the French National Research Infrastructure France-BioImaging (ANR10-INBS-04). This work was supported by Human Frontier Science Program (HFSP) grant RGP0005/2016 (to F.-C.T., J.M.H., G.A.V., P.L., and P.B.), Institut Curie and the Centre National de la Recherche Scientifique (CNRS) (to F.-C.T., J.M.H., and P.B.), Marie Curie actions H2020-MSCA-IF-2014 (to F.-C.T.), EMBO Long-Term fellowship ALTF 1527-2014 (to F.-C.T.), Pasteur Foundation Fellowship (to J.M.H.), Agence Nationale pour la Recherche ANR-20-CE13-0032 (to J.M.H. and P.B.) and ANR-20-CE11-0010-01 (to F.-C.T), Université Paris Sciences et Lettres-QLife Institute ANR-17-CONV-0005 Q-LIFE (to P.B.), FY 2015 Researcher Exchange Program between the Japan Society for the Promotion of Science and Academy of Finland (to Y.S.), the Takeda Science Foundation (to Y.S.), the Wesco Scientific Promotion Foundation (to Y.S.), Agence Nationale pour la Recherche ANR-18-CE13-0026-01 and ANR-21-CE13-0010-03 (to C.L.C.), Cancer Society Finland 4705949 (to P.L.), and U.S. National Institutes of Health (NIH) Institute of General Medical Sciences (NIGMS) grant R01-GM063796 (to G.A.V. and Z.J.) Publisher Copyright: Copyright © 2022 The Authors, some rights reserved. Filopodia are actin-rich membrane protrusions essential for cell morphogenesis, motility, and cancer invasion. How cells control filopodium initiation on the plasma membrane remains elusive. We performed experiments in cellulo, in vitro, and in silico to unravel the mechanism of filopodium initiation driven by the membrane curvature sensor IRSp53 (insulin receptor substrate protein of 53 kDa). We showed that full-length IRSp53 self-assembles into clusters on membranes depending on PIP2. Using well-controlled in vitro reconstitution systems, we demonstrated that IRSp53 clusters recruit the actin polymerase VASP (vasodilator-stimulated phosphoprotein) to assemble actin filaments locally on membranes, leading to the generation of actin-filled membrane protrusions reminiscent of filopodia. By pulling membrane nanotubes from live cells, we observed that IRSp53 can only be enriched and trigger actin assembly in nanotubes at highly dynamic membrane regions. Our work supports a regulation mechanism of IRSp53 in its attributes of curvature sensation and partner recruitment to ensure a precise spatial-temporal control of filopodium initiation.

(1) Hyperglycemia and oral pathology accelerate each other in diabetes. We evaluated whether gestational diabetes mellitus (GDM) is associated with self-reported increased oral health care needs and oral symptoms, including third molar symptoms, during pregnancy. (2) Pregnant women with (n = 1030) and without GDM (n = 935) were recruited in this multicenter Finnish Gestational Diabetes study in 2009–2012. Of the women with GDM, 196 (19.0%) receiving pharmacological treatment, 797 (77.0%) receiving diet treatment and 233 (23.0%) with recurrent GDM were analyzed separately. Oral health was assessed using structured questionnaires and analyzed by multivariable logistic regression adjusted for background risk factors. (3) Women with GDM were more likely to report a higher need for oral care than controls (31.1% vs. 24.5%; odds ratio (OR) 1.39; 95% confidence interval (CI) 1.14–1.69), particularly women with recurrent GDM (38.1% vs. 24.5%; OR 1.90; 95% CI 1.40–2.58). Women with pharmacologically treated GDM (46.9%) more often had third molar symptoms than controls (36.1%; OR 1.57; 95% CI 1.15–2.15) than women with diet-treated GDM (38.0%; OR 1.47; 95% CI 1.07–2.02). (4) GDM is associated with perceived oral care needs. Third molar symptoms were associated with pharmacologically treated GDM. publishedVersion

The study aimed to clarify psychiatric morbidity in patients who underwent orthognathic surgery (OS) pre- and postoperatively.Patients ≥18 years undergoing OS were included in this retrospective study. The outcome variable was the incidence of new mild, moderate, or severe psychiatric morbidity or exacerbation of preexisting psychiatric morbidity postoperatively. Surgery and patient-related background variables for outcome were analyzed (SPSS for Macintosh, version 27; IBM SPSS, Inc., Armonk, NY, USA).Of 182 patients, 44 (24%) had preceding psychiatric morbidity. It was associated significantly with history of alcohol abuse (P.001) and smoking (P = .046) and was more common in older patients (P = .042). During the postoperative phase, new psychiatric morbidity or exacerbation of a preexisting psychiatric condition was found in 12 patients (7%). Preceding psychiatric history (OR 8.88, P = .004) and high-dose perioperative dexamethasone (OR 9.81, P = .036) were independent predictors for postoperative psychiatric morbidity. No other evaluated variables were associated with outcome.Psychiatric conditions are common among OS patients. Treatment planning should consider the patient's mental health to minimize the risk of exacerbating psychiatric conditions, and collaboration with psychiatric professionals is recommended. Perioperative high-dose dexamethasone should be used with caution considering possible adverse psychiatric effects.

Polymerization of actin filaments against membranes produces force for numerous cellular processes, such as migration, morphogenesis, endocytosis, phagocytosis and organelle dynamics. Consequently, aberrant actin cytoskeleton dynamics are linked to various diseases, including cancer, as well as immunological and neurological disorders. Understanding how actin filaments generate forces in cells, how force production is regulated by the interplay between actin-binding proteins and how the actin-regulatory machinery responds to mechanical load are at the heart of many cellular, developmental and pathological processes. During the past few years, our understanding of the mechanisms controlling actin filament assembly and disassembly has evolved substantially. It has also become evident that the activities of key actin-binding proteins are not regulated solely by biochemical signalling pathways, as mechanical regulation is critical for these proteins. Indeed, the architecture and dynamics of the actin cytoskeleton are directly tuned by mechanical load. Here we discuss the general mechanisms by which key actin regulators, often in synergy with each other, control actin filament assembly, disassembly, and monomer recycling. By using an updated view of actin dynamics as a framework, we discuss how the mechanics and geometry of actin networks control actin-binding proteins, and how this translates into force production in endocytosis and mesenchymal cell migration.

Metastasis-suppressor 1 (MTSS1) is a membrane-interacting scaffolding protein that regulates the integrity of epithelial cell-cell junctions and functions as a tumor suppressor in a wide range of carcinomas. MTSS1 binds phosphoinositide-rich membranes through its I-BAR domain and is capable of sensing and generating negative membrane curvature in vitro. However, the mechanisms by which MTSS1 localizes to inter-cellular junctions in epithelial cells and contributes to their integrity and maintenance have remained elusive. By carrying out EM and live-cell imaging on cultured Madin-Darby canine kidney cell monolayers, we provide evidence that adherens junctions of epithelial cells harbor lamellipodia-like, dynamic actin-driven membrane folds, which exhibit high negative membrane curvature at their distal edges. BioID proteomics and imaging experiments demonstrated that MTSS1 associates with an Arp2/3 complex activator, the WAVE-2 complex, in dynamic actin-rich protrusions at cell-cell junctions. Inhibi-tion of Arp2/3 or WAVE-2 suppressed actin filament assembly at adherens junctions, decreased the dynamics of junctional membrane protrusions, and led to defects in epithelial integ-rity. Together, these results support a model in which membrane-associated MTSS1, together with the WAVE-2 and Arp2/3 complexes, promotes the formation of dynamic lamellipodia-like actin protrusions that contribute to the integrity of cell-cell junctions in epithelial monolayers.

Actin polymerization provides force for vital processes of the eukaryotic cell, but our understanding of actin dynamics and energetics remains limited due to the lack of high-quality probes. Most current probes affect dynamics of actin or its interactions with actin-binding proteins (ABPs), and cannot track the bound nucleotide. Here, we identify a family of highly sensitive fluorescent nucleotide analogues structurally compatible with actin. We demonstrate that these fluorescent nucleotides bind to actin, maintain functional interactions with a number of essential ABPs, are hydrolyzed within actin filaments, and provide energy to power actin-based processes. These probes also enable monitoring actin assembly and nucleotide exchange with single-molecule microscopy and fluorescence anisotropy kinetics, therefore providing robust and highly versatile tools to study actin dynamics and functions of ABPs., Actin polymerization provides force for vital processes of the eukaryotic cell, but our understanding of actin dynamics and energetics remains limited due to the lack of high-quality probes. Here authors identify a family of highly sensitive fluorescent nucleotide analogues which bind to actin and provide energy to power actin-based processes.

The increased use of social media in dentistry is associated with both advantages and disadvantages. A new form of professionalism, “e-professionalism,” has emerged. It includes an online persona and online information in any format that displays cues to professional identity, attitudes, and behaviors. The objective was to explore the perceptions of Malaysian and Finnish dental students on e-professionalism. A survey of 613 Malaysian and Finnish students was performed. The main variables assessed were posting of objectionable or inappropriate content among students, attitudes towards unprofessional online content, perceived online presence, contacts with patients and faculty members on social media, and concerns about social media use. The prevalence of posting clearly unprofessional content was not high among dental students. Revealing information of patients was most common content of clear unprofessionalism. Students from Malaysia contacted patients and faculty members more actively in social media than students in Finland (73.6% of students in Malaysia and 11.8% in Finland had invited faculty members to be “friends”). Malaysian students were more concerned and more likely to react to inappropriate content on social media. Attitude of dental students towards social media use in dentistry were very positive in both countries. Students agreed that guiding patients online is a new responsibility for dentists in the digital age (86.4% of students in Malaysia and 73.4% in Finland). The findings indicate the existence of both benefits and dangers of social media on e-professionalism among students. There is a need to include robust digital professionalism awareness training for students.

Contractile actomyosin bundles, stress fibers, contribute to morphogenesis, migration, and mechanosensing of non-muscle cells. In addition to actin and non-muscle myosin II (NMII), stress fibers contain a large array of proteins that control their assembly, turnover, and contractility. Calponin-3 (Cnn3) is an actin-binding protein that associates with stress fibers. However, whether Cnn3 promotes stress fiber assembly, or serves as either a positive or negative regulator of their contractility has remained obscure. Here, we applied U2OS osteosarcoma cells as a model system to study the function of Cnn3. We show that Cnn3 localizes to both NMII-containing contractile ventral stress fibers and transverse arcs, as well as to non-contractile dorsal stress fibers that do not contain NMII. Fluorescence-recovery-after-photobleaching experiments revealed that Cnn3 is a dynamic component of stress fibers. Importantly, CRISPR/Cas9 knockout and RNAi knockdown studies demonstrated that Cnn3 is not essential for stress fiber assembly. However, Cnn3 depletion resulted in increased and uncoordinated contractility of stress fibers that often led to breakage of individual actomyosin bundles within the stress fiber network. Collectively these results provide evidence that Cnn3 is dispensable for the assembly of actomyosin bundles, but that it is required for controlling proper contractility of the stress fiber network.

The cerebral cortex is responsible for higher cognitive functions. Its correct development depends on coordinated asymmetric division cycles by polarized radial glial progenitor cells. Mitotic defects in neuronal stem cells are linked to the aetiology of microcephaly, but the underlying mechanisms remain incompletely understood. Here we describe a novel role of the membrane deforming cytoskeletal regulator protein Abba (MTSS1L/MTSS2) in cortical development. Loss of Abba in the developing brain resulted in radial glia proliferation arrest, disorganized radial fibers and abnormal migration of neuronal progenitors. During cell division, Abba localized to the cleavage furrow where it recruited the scaffolding protein Nedd9 and positively regulated RhoA activity. Importantly, we identified a human variant Abba (R671W) in a patient with microcephaly and intellectual disability. Ectopic expression of this mutant protein in mice phenocopied Abba knockdown. Taken together, these data provide novel mechanistic insight into human microcephaly and cortical development by identifying ABBA as a novel regulator of faithful mitotic progression of neuronal progenitor cells.

Author summaryRelapsing fever and Lyme Disease are infectious diseases caused by borrelia bacteria. Relapsing fever occurs sporadically worldwide, whereas distribution of Lyme Disease is restricted to the Northern Hemisphere. Both infections are transmitted to humans by blood eating ticks or lice. These infections are often difficult to diagnose due to nonspecific symptoms. To be able to cause infection, borrelia must circumvent the human immune responses. Here we describe a mechanism, how borrelia bacteria protect themselves in the human host by utilizing host proteins. By using X-ray crystallography, we solved the structure of an outer membrane protein FhbA from a relapsing fever causing borreliae, Borrelia hermsii, in complex with human complement regulator factor H. FhbA has a unique alpha-helical fold that has not been reported earlier. The structure of the complex revealed how FhbA binds factor H in a very specific manner. Factor H bound to FhbA on the surface of borrelia protects bacteria from the complement system and lysis. Based on the structure, we performed structure-guided sequence database analysis, which suggests that similar proteins are present in all relapsing fever causing borrelia and possibly in some Lyme disease agents. Immune evasion facilitates survival of Borrelia, leading to infections like relapsing fever and Lyme disease. Important mechanism for complement evasion is acquisition of the main host complement inhibitor, factor H (FH). By determining the 2.2 angstrom crystal structure of Factor H binding protein A (FhbA) from Borrelia hermsii in complex with FH domains 19-20, combined with extensive mutagenesis, we identified the structural mechanism by which B. hermsii utilizes FhbA in immune evasion. Moreover, structure-guided sequence database analysis identified a new family of FhbA-related immune evasion molecules from Lyme disease and relapsing fever Borrelia. Conserved FH-binding mechanism within the FhbA-family was verified by analysis of a novel FH-binding protein from B. duttonii. By sequence analysis, we were able to group FH-binding proteins of Borrelia into four distinct phyletic types and identified novel putative FH-binding proteins. The conserved FH-binding mechanism of the FhbA-related proteins could aid in developing new approaches to inhibit virulence and complement resistance in Borrelia.

A large proportion of proteins are expected to interact with cellular membranes to carry out their physiological functions in processes such as membrane transport, morphogenesis, cytoskeletal organization, and signal transduction. The recruitment of proteins at the membrane-cytoplasm interface and their activities are precisely regulated by phosphoinositides, which are negatively charged phospholipids found on the cytoplasmic leaflet of cellular membranes and play critical roles in membrane homeostasis and cellular signaling. Thus, it is important to reveal which proteins interact with phosphoinositides and to elucidate the underlying mechanisms. Here, we present two standard in vitro methods, liposome co-sedimentation and co-flotation assays, to study lipid-protein interactions. Liposomes can mimic various biological membranes in these assays because their lipid compositions and concentrations can be varied. Thus, in addition to mechanisms of lipid-protein interactions, these methods provide information on the possible specificities of proteins toward certain lipids such as specific phosphoinositide species and can hence shed light on the roles of membrane interactions on the functions of membrane-associated proteins.

The actomyosin cytoskeleton serves as a key regulator of the integrity and remodeling of epithelial barriers by controlling assembly and functions of intercellular junctions and cell-matrix adhesions. While biochemical mechanisms that regulate activity of non-muscle myosin II (NM-II) in epithelial cells have been extensively investigated, little is known about assembly of the contractile myosin structures at the epithelial adhesion sites. UNC-45A is a cytoskeletal chaperone that is essential for proper folding of NM II heavy chains and myofilament assembly. We found abundant expression of UNC-45A in human intestinal epithelial cell (IEC) lines and in the epithelial layer of the normal human colon. Interestingly, protein levels of UNC-45A were decreased in colonic epithelium of patients with ulcerative colitis. CRISPR/Cas9-mediated knock-out of UNC-45A in HT-29 and SK-CO15 IEC disrupted epithelial barrier integrity, impaired assembly of epithelial adherence and tight junctions and attenuated cell migration. Consistently, decreased UNC-45 expression increased permeability of the Drosophila gut in vivo. The mechanisms underlying barrier disruptive and anti-migratory effects of UNC-45A depletion involved disorganization of the actomyosin bundles at epithelial junctions and the migrating cell edge. Loss of UNC-45A also decreased contractile forces at epithelial junctions and matrix adhesions. Expression of deletion mutants revealed roles for the myosin binding domain of UNC-45A in controlling IEC junctions and motility. Our findings uncover a novel mechanism that regulates integrity and restitution of the intestinal epithelial barrier, which may be impaired during mucosal inflammation.

Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. Social media use among students has infiltrated into dental education and offers benefits but may also cause problems. The aim of this study was to explore and compare current social media usage among dental undergraduate students from two countries—Malaysia and Finland. A self-administered structured online questionnaire was used. WhatsApp, YouTube, Instagram, Facebook and Snapchat were the services that were most familiar to the respondents from both countries. There were differences between the students from the two countries among the most preferred platforms. The most frequently used applications were WhatsApp (91.1% of students in Malaysia and 96.1% in Finland used it very frequently) and Instagram (74.3% of students in Malaysia and 70.0% in Finland used it very frequently). Students in Malaysia spent significantly more hours per week using the platforms as study tools than students in Finland. Over 80% of the Finnish dental students reported that lack of knowledge was not an issue in social media usage, while 85% of Malaysian students felt that lack of knowledge prevented them from using social media platforms frequently. The findings offer evidence that dental students used social media extensively.

Eukaryotic cells have diverse protrusive and contractile actin filament structures, which compete with one another for a limited pool of actin monomers. Numerous actin-binding proteins regulate the dynamics of actin structures, including tropomodulins (Tmods), which cap the pointed end of actin filaments. In striated muscles, Tmods prevent actin filaments from overgrowing, whereas in non-muscle cells, their function has remained elusive. Here, we identify two Tmod isoforms, Tmod1 and Tmod3, as key components of contractile stress fibers in non-muscle cells. Individually, Tmodl and Tmod3 can compensate for one another, but their simultaneous depletion results in disassembly of actin-tropomyosin filaments, loss of force-generating stress fibers, and severe defects in cell morphology. Knockout-rescue experiments reveal that Tmod's interaction with tropomyosin is essential for its role in the stabilization of actin-tropo-myosin filaments in cells. Thus, in contrast to their role in muscle myofibrils, in non-muscle cells, Tmods bind actin-tropomyosin filaments to protect them from depolymerizing, not elongating. Furthermore, loss of Tmods shifts the balance from linear actin-tropomyosin filaments to Arp2/3 complex-nucleated branched networks, and this phenotype can be partially rescued by inhibiting the Arp2/3 complex. Collectively, the data reveal that Tmods are essential for the maintenance of contractile actomyosin bundles and that Tmod-dependent capping of actin-tropomyosin filaments is critical for the regulation of actin homeostasis in non-muscle cells.

Actin polymerization powers key cellular processes, including motility, morphogenesis, and endocytosis. The actin turnover cycle depends critically on “re-charging” of ADP-actin monomers with ATP, but whether this reaction requires dedicated proteins in cells, and the underlying mechanism, have remained elusive. Here we report that nucleotide exchange catalyzed by the ubiquitous cytoskeletal regulator cyclase-associated protein (CAP) is critical for actin-based processes in vivo. We determine the structure of the CAP–actin complex, which reveals that nucleotide exchange occurs in a compact, sandwich-like complex formed between the dimeric actin-binding domain of CAP and two ADP-actin monomers. In the crystal structure, the C-terminal tail of CAP associates with the nucleotide-sensing region of actin, and this interaction is required for rapid re-charging of actin by both yeast and mammalian CAPs. These data uncover the conserved structural basis and biological role of protein-catalyzed re-charging of actin monomers., Depolymerized ADP-actin monomers must be recharged with ATP for new rounds of filament assembly. Here the authors show that cyclase-associated protein (CAP) catalyzes actin nucleotide exchange in vivo and their CAP–actin complex structure reveals the molecular mechanism of CAP-mediated actin nucleotide exchange.

International audience; During HIV-1 particle formation, the requisite plasma membrane curvature is thought to be solely driven by the retroviral Gag protein. Here, we reveal that the cellular I-BAR protein IRSp53 is required for the progression of HIV-1 membrane curvature to complete particle assembly. siRNA-mediated knockdown of IRSp53 gene expression induces a decrease in viral particle production and a viral bud arrest at half completion. Single-molecule localization microscopy at the cell plasma membrane shows a preferential localization of IRSp53 around HIV-1 Gag assembly sites. In addition, we observe the presence of IRSp53 in purified HIV-1 particles. Finally, HIV-1 Gag protein preferentially localizes to curved membranes induced by IRSp53 I-BAR domain on giant unilamellar vesicles. Overall, our data reveal a strong interplay between IRSp53 I-BAR and Gag at membranes during virus assembly. This highlights IRSp53 as a crucial host factor in HIV-1 membrane curvature and its requirement for full HIV-1 particle assembly.

The actin cytoskeleton is indispensable for several cellular processes, including migration, morphogenesis, polarized growth, endocytosis, and phagocytosis. The organization and dynamics of the actin cytoskeleton in these processes are regulated by Rho family small GTPases and kinase-phosphatase pathways. Moreover, membrane phospholipids, especially the phosphatidylinositol phosphates have emerged as important regulators of actin dynamics. From these, PI(4,5)P2 is the most abundant at the plasma membrane, and directly regulates the activities and subcellular localizations of numerous actin-binding proteins. Here, we discuss recent studies demonstrating that actin-binding proteins interact with PI(4,5)P2-rich membranes through drastically different affinities and dynamics correlating with their roles in cytoskeletal dynamics. Moreover, by using mesenchymal cell migration and clathrin-mediated endocytosis as examples, we present a model for how interplay between PI(4,5)P2 and actin-binding proteins control the actin cytoskeleton in cells.