The small molecule inhibitor NAV-2729 has a complex target profile including multiple ADP-ribosylation factor regulatory proteins.

The ADP-ribosylation factor (Arf) GTPases and their regulatory proteins are implicated in cancer progression. NAV-2729 was previously identified as a specific inhibitor of Arf6 that reduced progression of uveal melanoma in an orthotopic xenograft. Here, our goal was to assess the inhibitory effects of NAV-2729 on the proliferation of additional cell types. We found NAV-2729 inhibited proliferation of multiple cell lines, but Arf6 expression did not correlate with NAV-2729 sensitivity, and knockdown of Arf6 affected neither cell viability nor sensitivity to NAV-2729. Furthermore, binding to native Arf6 was not detected; however, we determined that NAV-2729 inhibited both Arf exchange factors and Arf GTPase-activating proteins. ASAP1, a GTPase-activating protein linked to cancer progression, was further investigated. We demonstrated that NAV-2729 bound to the PH domain of ASAP1 and changed ASAP1 cellular distribution. However, ASAP1 knockdown did not fully recapitulate the cytoskeletal effects of NAV-2729 nor affect cell proliferation. Finally, our screens identified 48 other possible targets of NAV-2729. These results illustrate the complexities of defining targets of small molecules and identify NAV-2729 as a model PH domain-binding inhibitor.

Collaborative Human-Computer Vision Operative Video Analysis Algorithm for Analyzing Surgical Fluency and Surgical Interruptions in Endonasal Endoscopic Pituitary Surgery: Cohort Study.

BACKGROUND: The endonasal endoscopic approach (EEA) is effective for pituitary adenoma resection. However, manual review of operative videos is time-consuming. The application of a computer vision (CV) algorithm could potentially reduce the time required for operative video review and facilitate the training of surgeons to overcome the learning curve of EEA. OBJECTIVE: This study aimed to evaluate the performance of a CV-based video analysis system, based on OpenCV algorithm, to detect surgical interruptions and analyze surgical fluency in EEA. The accuracy of the CV-based video analysis was investigated, and the time required for operative video review using CV-based analysis was compared to that of manual review. METHODS: The dominant color of each frame in the EEA video was determined using OpenCV. We developed an algorithm to identify events of surgical interruption if the alterations in the dominant color pixels reached certain thresholds. The thresholds were determined by training the current algorithm using EEA videos. The accuracy of the CV analysis was determined by manual review, and the time spent was reported. RESULTS: A total of 46 EEA operative videos were analyzed, with 93.6%, 95.1%, and 93.3% accuracies in the training, test 1, and test 2 data sets, respectively. Compared with manual review, CV-based analysis reduced the time required for operative video review by 86% (manual review: 166.8 and CV analysis: 22.6 minutes; P<.001). The application of a human-computer collaborative strategy increased the overall accuracy to 98.5%, with a 74% reduction in the review time (manual review: 166.8 and human-CV collaboration: 43.4 minutes; P<.001). Analysis of the different surgical phases showed that the sellar phase had the lowest frequency (nasal phase: 14.9, sphenoidal phase: 15.9, and sellar phase: 4.9 interruptions/10 minutes; P<.001) and duration (nasal phase: 67.4, sphenoidal phase: 77.9, and sellar phase: 31.1 seconds/10 minutes; P<.001) of surgical interruptions. A comparison of the early and late EEA videos showed that increased surgical experience was associated with a decreased number (early: 4.9 and late: 2.9 interruptions/10 minutes; P=.03) and duration (early: 41.1 and late: 19.8 seconds/10 minutes; P=.02) of surgical interruptions during the sellar phase. CONCLUSIONS: CV-based analysis had a 93% to 98% accuracy in detecting the number, frequency, and duration of surgical interruptions occurring during EEA. Moreover, CV-based analysis reduced the time required to analyze the surgical fluency in EEA videos compared to manual review. The application of CV can facilitate the training of surgeons to overcome the learning curve of endoscopic skull base surgery. TRIAL REGISTRATION: ClinicalTrials.gov NCT06156020; https://clinicaltrials.gov/study/NCT06156020.

The BAR domain of the Arf GTPase-activating protein ASAP1 directly binds actin filaments.

The Arf GTPase-activating protein (Arf GAP) with SH3 domain, ankyrin repeat and PH domain 1 (ASAP1) establishes a connection between the cell membrane and the cortical actin cytoskeleton. The formation, maintenance, and turnover of actin filaments and bundles in the actin cortex are important for cell adhesion, invasion, and migration. Here, using actin cosedimentation, polymerization, and depolymerization assays, along with total internal reflection fluorescence (TIRF), confocal, and EM analyses, we show that the N-terminal N-BAR domain of ASAP1 directly binds to F-actin. We found that ASAP1 homodimerization aligns F-actin in predominantly unipolar bundles and stabilizes them against depolymerization. Furthermore, the ASAP1 N-BAR domain moderately reduced the spontaneous polymerization of G-actin. The overexpression of the ASAP1 BAR-PH tandem domain in fibroblasts induced the formation of actin-filled projections more effectively than did full-length ASAP1. An ASAP1 construct that lacked the N-BAR domain failed to induce cellular projections. Our results suggest that ASAP1 regulates the dynamics and the formation of higher-order actin structures, possibly through direct binding to F-actin via its N-BAR domain. We propose that ASAP1 is a hub protein for dynamic protein-protein interactions in mechanosensitive structures, such as focal adhesions, invadopodia, and podosomes, that are directly implicated in oncogenic events. The effect of ASAP1 on actin dynamics puts a spotlight on its function as a central signaling molecule that regulates the dynamics of the actin cytoskeleton by transmitting signals from the plasma membrane.

ARAP2 inhibits Akt independently of its effects on focal adhesions.

BACKGROUND INFORMATION: ARAP2, an Arf GTPase-activating protein (Arf GAP) that binds to adaptor protein with PH domain, PTB domain and leucine zipper motifs 1 (APPL1), regulates focal adhesions (FAs). APPL1 affects FA dynamics by regulating Akt. Here, we tested the hypothesis that ARAP2 affects FAs in part by regulating Akt through APPL1. RESULTS: We found that ARAP2 controlled FA dynamics dependent on its enzymatic Arf GAP activity. In some cells, ARAP2 also regulated phosphoAkt (pAkt) levels. However, ARAP2 control of FAs did not require Akt and conversely, the effects on pAkt were independent of FAs. Reducing ARAP2 expression reduced the size and number of FAs in U118, HeLa and MDA-MB-231 cells. Decreasing ARAP2 expression increased pAkt in U118 cells and HeLa cells and overexpressing ARAP2 decreased pAkt in U118 cells; in contrast, ARAP2 had no effect on pAkt in MDA-MB-231 cells. An Akt inhibitor did not block the effect of reduced ARAP2 on FAs in U118. Furthermore, the effect of ARAP2 on Akt did not require Arf GAP activity, which is necessary for effects on FAs and integrin traffic. Altering FAs by other means did not induce the same changes in pAkt as those seen by reducing ARAP2 in U118 cells. In addition, we discovered that ARAP2 and APPL1 had co-ordinated effects on pAkt in U118 cells. Reduced APPL1 expression, as for ARAP2, increased pAkt in U118 and the effect of reduced APPL1 expression was reversed by overexpressing ARAP2. Conversely, the effect of reduced ARAP2 expression was reversed by overexpressing APPL1. ARAP2 is an Arf GAP that has previously been reported to affect FAs by regulating Arf6 and integrin trafficking and to bind to the adaptor proteins APPL1. Here, we report that ARAP2 suppresses pAkt levels in cells co-ordinately with APPL1 and independently of GAP activity and its effect on the dynamic behaviour of FAs. CONCLUSIONS: We conclude that ARAP2 affects Akt signalling in some cells by a mechanism independent of FAs or membrane traffic. SIGNIFICANCE: Our results highlight an Arf GAP-independent function of ARAP2 in regulating Akt activity and distinguish the effect of ARAP2 on Akt from that on FAs and integrin trafficking, which requires regulation of Arf6.

Arf GAPs as Regulators of the Actin Cytoskeleton-An Update.

Arf GTPase-activating proteins (Arf GAPs) control the activity of ADP-ribosylation factors (Arfs) by inducing GTP hydrolysis and participate in a diverse array of cellular functions both through mechanisms that are dependent on and independent of their Arf GAP activity. A number of these functions hinge on the remodeling of actin filaments. Accordingly, some of the effects exerted by Arf GAPs involve proteins known to engage in regulation of the actin dynamics and architecture, such as Rho family proteins and nonmuscle myosin 2. Circular dorsal ruffles (CDRs), podosomes, invadopodia, lamellipodia, stress fibers and focal adhesions are among the actin-based structures regulated by Arf GAPs. Arf GAPs are thus important actors in broad functions like adhesion and motility, as well as the specialized functions of bone resorption, neurite outgrowth, and pathogen internalization by immune cells. Arf GAPs, with their multiple protein-protein interactions, membrane-binding domains and sites for post-translational modification, are good candidates for linking the changes in actin to the membrane. The findings discussed depict a family of proteins with a critical role in regulating actin dynamics to enable proper cell function.

Direct Functional Interaction of the Kinesin-13 Family Member Kinesin-like Protein 2A (Kif2A) and Arf GAP with GTP-binding Protein-like, Ankyrin Repeats and PH Domains1 (AGAP1).

The molecular basis for control of the cytoskeleton by the Arf GTPase-activating protein AGAP1 has not been characterized. AGAP1 is composed of G-protein-like (GLD), pleckstrin homology (PH), Arf GAP, and ankyrin repeat domains. Kif2A was identified in screens for proteins that bind to AGAP1. The GLD and PH domains of AGAP1 bound the motor domain of Kif2A. Kif2A increased GAP activity of AGAP1, and a protein composed of the GLD and PH domains of AGAP1 increased ATPase activity of Kif2A. Knockdown (KD) of Kif2A or AGAP1 slowed cell migration and accelerated cell spreading. The effect of Kif2A KD on spreading could be rescued by expression of Kif2A-GFP or FLAG-AGAP1, but not by Kif2C-GFP. The effect of AGAP1 KD could be rescued by FLAG-AGAP1, but not by an AGAP1 mutant that did not bind Kif2A efficiently, ArfGAP1-HA or Kif2A-GFP. Taken together, the results support the hypothesis that the Kif2A·AGAP1 complex contributes to control of cytoskeleton remodeling involved in cell movement.

The Arf GTPase-activating Protein, ASAP1, Binds Nonmuscle Myosin 2A to Control Remodeling of the Actomyosin Network.

ASAP1 regulates F-actin-based structures and functions, including focal adhesions (FAs) and circular dorsal ruffles (CDRs), cell spreading and migration. ASAP1 function requires its N-terminal BAR domain. We discovered that nonmuscle myosin 2A (NM2A) directly bound the BAR-PH tandem of ASAP1in vitro ASAP1 and NM2A co-immunoprecipitated and colocalized in cells. Knockdown of ASAP1 reduced colocalization of NM2A and F-actin in cells. Knockdown of ASAP1 or NM2A recapitulated each other's effects on FAs, cell migration, cell spreading, and CDRs. The NM2A-interacting BAR domain contributed to ASAP1 control of cell spreading and CDRs. Exogenous expression of NM2A rescued the effect of ASAP1 knockdown on CDRs but ASAP1 did not rescue NM2A knockdown defect in CDRs. Our results support the hypothesis that ASAP1 is a positive regulator of NM2A. Given other binding partners of ASAP1, ASAP1 may directly link signaling and the mechanical machinery of cell migration.

The Arf6 GTPase-activating proteins ARAP2 and ACAP1 define distinct endosomal compartments that regulate integrin α5ß1 traffic.

Arf6 and the Arf6 GTPase-activating protein (GAP) ACAP1 are established regulators of integrin traffic important to cell adhesion and migration. However, the function of Arf6 with ACAP1 cannot explain the range of Arf6 effects on integrin-based structures. We propose that Arf6 has different functions determined, in part, by the associated Arf GAP. We tested this idea by comparing the Arf6 GAPs ARAP2 and ACAP1. We found that ARAP2 and ACAP1 had opposing effects on apparent integrin ß1 internalization. ARAP2 knockdown slowed, whereas ACAP1 knockdown accelerated, integrin ß1 internalization. Integrin ß1 association with adaptor protein containing a pleckstrin homology (PH) domain, phosphotyrosine-binding (PTB) domain, and leucine zipper motif (APPL)-positive endosomes and EEA1-positive endosomes was affected by ARAP2 knockdown and depended on ARAP2 GAP activity. ARAP2 formed a complex with APPL1 and colocalized with Arf6 and APPL in a compartment distinct from the Arf6/ACAP1 tubular recycling endosome. In addition, although ACAP1 and ARAP2 each colocalized with Arf6, they did not colocalize with each other and had opposing effects on focal adhesions (FAs). ARAP2 overexpression promoted large FAs, but ACAP1 overexpression reduced FAs. Taken together, the data support a model in which Arf6 has at least two sites of opposing action defined by distinct Arf6 GAPs.

Sandensone A, a novel sesquiterpenoid from the Formosan soft coral Sinularia sandensis.

A chromatographic and NMR spectroscopy-based fractionation on the acetone extracts of the soft coral Sinularia sandensis led to the isolation of a novel sesquiterpenoid, sandensone A (1). The structure of 1 was elucidated based on comprehensive 1D and 2D NMR spectroscopic data as well as HRESIMS spectrometry. The absolute configuration at C-12 of 1 was determined as R using a modified Mosher's method. The cytotoxicity against A549 (human lung carcinoma), HT-29 (human colon adenocarcinoma), and P-388 (mouse lymphocytic leukemia) cancer cell lines as well as antiviral activity against HCMV (human cytomegalovirus) were evaluated in vitro.

Ultra-structure changes and survivin expression in uterine fibroids after radiofrequency ablation.

PURPOSE: To explore the reliability and validity of radiofrequency (RF) ablation in treating uterine fibroids. MATERIALS AND METHODS: We evaluated 63 patients who underwent hysterectomy to treat multiple fibroids. Thirty patients immediately underwent abdominal hysterectomy after the fibroids were ablated under direct vision. Thirty-three patients first experienced trans-vaginal ablation with the guidance of a baseline ultrasound. We performed abdominal or trans-vaginal hysterectomy 72 h later. The tissues in the centre of the ablated lesion (group A), at the edge of the ablated lesion (group B), 1 cm away from the ablated edge (group C) and the control group were sampled. We observed ultra-structure changes by transmission electron microscopy and detected survivin expression with Western blot analysis. RESULTS: According to transmission electron microscopy, the ultra-structure of fibroid cells in groups A and B was damaged. However, in group C, the ultra-structure was normal. Compared with the control group, survivin expression was significantly decreased. Meanwhile survivin expression was significantly increased with the distance to the ablated centre (p < 0.05). CONCLUSIONS: Radiofrequency ablation caused permanent and irreversible damage to fibroid cells and decreased survivin expression, which provided reliable clinical evidence for the success of radiofrequency ablation treating uterine fibroids.

ARAP2 signals through Arf6 and Rac1 to control focal adhesion morphology.

Focal adhesions (FAs) are dynamic structures that connect the actin cytoskeleton with the extracellular matrix. At least six ADP-ribosylation factor (Arf) GTPase-activating proteins (GAPs), including ARAP2 (an Arf6 GAP), are implicated in regulation of FAs but the mechanisms for most are not well defined. Although Rac1 has been reported to function downstream of Arf6 to control membrane ruffling and cell migration, this pathway has not been directly examined as a regulator of FAs. Here we test the hypothesis that ARAP2 promotes the growth of FAs by converting Arf6·GTP to Arf6·GDP thereby preventing the activation of the Rho family GTP-binding protein Rac1. Reduced expression of ARAP2 decreased the number and size of FAs in cells and increased cellular Arf6·GTP and Rac1·GTP levels. Overexpression of ARAP2 had the opposite effects. The effects of ARAP2 on FAs and Rac1 were dependent on a functional ArfGAP domain. Constitutively active Arf6 affected FAs in the same way as did reduced ARAP2 expression and dominant negative mutants of Arf6 and Rac1 reversed the effect of reduced ARAP2 expression. However, neither dominant negative Arf6 nor Rac1 had the same effect as ARAP2 overexpression. We conclude that changes in Arf6 and Rac1 activities are necessary but not sufficient for ARAP2 to promote the growth of FAs and we speculate that ARAP2 has additional functions that are effector in nature to promote or stabilize FAs.

Control of cell signaling by Arf GTPases and their regulators: Focus on links to cancer and other GTPase families.

The ADP-ribosylation factors (Arfs) comprise a family of regulatory GTP binding proteins. The Arfs regulate membrane trafficking and cytoskeleton remodeling, processes critical for eukaryotes and which have been the focus of most studies on Arfs. A more limited literature describes a role in signaling and in integrating several signaling pathways to bring about specific cell behaviors. Here, we will highlight work describing function of Arf1, Arf6 and several effectors and regulators of Arfs in signaling.

ERK phosphorylation is dependent on cell adhesion in a subset of pediatric sarcoma cell lines.

Osteosarcoma (OS) and Pax-Foxo1 fusion negative rhabdomyosarcoma (FN-RMS) are pediatric sarcomas with poor prognoses in patients with advanced disease. In both malignancies, an actin binding protein has been linked to poor prognosis. Integrin adhesion complexes (IACs) are closely coupled to actin networks and IAC-mediated signaling has been implicated in the progression of carcinomas. However, the relationship of IACs and actin cytoskeleton remodeling with cell signaling is understudied in pediatric sarcomas. Here, we tested the hypothesis that IAC dynamics affect ERK activation in OS and FN-RMS cell lines. Adhesion dependence of ERK activation differed among the OS and FN-RMS cells examined. In the OS cell lines, adhesion did not have a consistent effect on phospho-ERK (pERK). ERK phosphorylation in response to fetal calf serum or 1 ng/ml EGF was nearly as efficient in OS cell lines and one FN-RMS cell line in suspension as cells adherent to poly-l-lysine (PL) or fibronectin (FN). By contrast, adhesion to plastic, PL or FN increased ERK phosphorylation and was greater than additive with a 15 min exposure to 1 ng/ml EGF in three FN-RMS cell lines. Increases in pERK were partly dependent on FAK and PAK1/2 but independent of IAC maturation. As far as we are aware, this examination of adhesion-dependent signaling is the first in pediatric sarcomas and has led to the discovery of differences from the prevailing paradigms and differences in the degree of coupling between components in the signaling pathways among the cell lines.

New cembranolides from the Dongsha Atoll soft coral Lobophytum durum.

Chemical investigations of the Dongsha Atoll soft coral Lobophytum durum resulted in the isolation of five new cembranolides, durumolides M-Q (1-5). The structures of compounds 1-5 were characterized by the interpretation of extensive spectroscopic analysis. Compound 4 exhibited cytotoxicity against P-388 (mouse lymphocytic leukemia) cell line with an ED50 of 3.8 µg/mL. Moreover, compound 5 showed significant antiviral activity against human cytomegalovirus with an IC50 of 5.2 µg/mL.

Coenzyme Q10 amends testicular function and spermatogenesis in male mice exposed to cigarette smoke by modulating oxidative stress and inflammation.

This study explored the effects of coenzyme Q10 (CoQ10) on the testicular functions of male mice exposed to cigarette smoke. Eight-week-old BALB/c male mice were divided into the following groups: the AV group (air with a vehicle), the AQ group (air with CoQ10), the SV group (smoke with a vehicle), and the SQ group (smoke with CoQ10). The results showed that the CoQ10 concentrations in the sera and testes were decreased in the groups subjected to smoke but they were improved after the administration of CoQ10. Neither smoke nor CoQ10 supplementation affected the serum or testis testosterone concentrations. Regarding the antioxidant system in the testis, the exposure to smoke induced malondialdehyde and hydrogen peroxide production and decreased the catalase and glutathione peroxidase activities. Oral CoQ10 administration reversed the oxidative damage. In apoptosis, the cytochrome c, c-caspase 9, and c-caspase 3 proteins were increased in the groups exposed to smoke but they were decreased after the CoQ10 administration. In mitochondrial biogenesis, smoke exposure led to decreases in the PGC1-α, NRF1, and NRF2 levels, but CoQ10 increased the expressions of these proteins. Additionally, oral CoQ10 administration improved the mitochondrial copy numbers that were reduced following the exposure to smoke. In summary, CoQ10 administration reduces smoke-induced testicular damage by regulating the antioxidant capacity, the cell apoptosis, the mitochondrial biogenesis, and the copy numbers in the testes.

Memantine Protects against Paclitaxel-Induced Cognitive Impairment through Modulation of Neurogenesis and Inflammation in Mice.

Chemotherapy-induced cognitive impairment (CICI) is an adverse side effect of cancer treatment with increasing awareness. Hippocampal damage and related neurocognitive impairment may mediate the development of CICI, in which altered neurogenesis may play a role. In addition, increased inflammation may be related to chemotherapy-induced hippocampal damage. Memantine, an uncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist that may enhance neurogenesis and modulate inflammation, may be useful for treating CICI. To test this hypothesis, paclitaxel was administered to eight-week-old male B6 mice to demonstrate the relationship between CICI and impaired neurogenesis, and then, we evaluated the impact of different memantine regimens on neurogenesis and inflammation in this CICI model. The results demonstrated that both the pretreatment and cotreatment regimens with memantine successfully reversed impaired neurogenesis and spatial memory impairment in behavior tests. The pretreatment regimen unsuccessfully inhibited the expression of peripheral and central TNF-α and IL-1ß and did not improve the mood alterations following paclitaxel treatment. However, the cotreatment regimen led to a better modulatory effect on inflammation and restoration of mood disturbance. In conclusion, this study illustrated that impaired neurogenesis is one of the mechanisms of paclitaxel-induced CICI. Memantine may serve as a potential treatment for paclitaxel-induced CICI, but different treatment strategies may lead to variations in the treatment efficacy.

Antiviral and anti-inflammatory metabolites from the soft coral Sinularia capillosa.

Chemical investigations of the soft coral Sinularia capillosa resulted in the isolation of one new tetraprenylbenzoquinone, capilloquinone (1), two new furanobenzosesquiterpenoids, capillobenzopyranol (2) and capillobenzofuranol (3), one new furanosesquiterpenoid, capillofuranocarboxylate (4), and five previously characterized metabolites, comprising (E)-5-(2,6-dimethylocta-5,7-dienyl)furan-3-carboxylic acid (5), 2-[(2E,6E)-3,7-dimethyl-8-(4-methylfuran-2-yl)octa-2,6-dienyl]-5-methylcyclohexa-2,5-diene-1,4-dione (6), 2-[(2E,6E)-3,7-dimethyl-8-(4-methylfuran-2-yl)octa-2,6-dienyl]-5-methylbenzene-1,4-diol (7), (-)-loliolide (8), and 3,4,11-trimethyl-7-methylenebicyclo[6.3.0]undec-2-en-11alpha-ol (9). The structures of 1-4 were elucidated through extensive spectroscopic analysis. The cytotoxicity, anti-HCMV (human cytomegalovirus) activity, antibacterial activity, and anti-inflammatory effects of 1-9 were evaluated in vitro.

Accumulation of gallium (Ga) and indium (In) in rice grains in Ga- and In-contaminated paddy soils.

To understand the risk of two emerging contaminants, gallium (Ga) and indium (In) to humans via rice consumption, effects of soil properties and concentrations of spiked Ga/In on the accumulation of Ga and In in rice grains were investigated. A pot experiment was conducted, and paddy rice was grown in three soils with different pH values and Al availabilities (i.e., Pc, TWz and Cf), which were spiked with various Ga and In concentrations. The growth index and concentrations of Ga, In, and Al in plant tissues and soil pore water were measured. Results revealed that the concentrations of Ga and In in soil pore water increase with the spiking of Ga or In in all of the tested soils, but the biomass of roots and shoots does not significantly decrease. The accumulation of Ga in shoots and brown rice was significantly reduced in high available Al acidic soils (Pc soils), but this accumulation was significantly increased in low available Al acidic soils (TWz soils), which can be explained by the competitive uptake between Ga and Al by rice plants. The extent of competitive effects between In and Al was less than that between Ga and Al because of the lower solubility and translocation capability of In than those of Ga in soil-rice systems. However, significant differences in the concentrations of Ga and In in brown rice in neutral soils (Cf soils) among the Ga or In treatment were not observed. In addition, the iron plaque formed on the root surface can serve as a barrier to reduce the accumulation of Ga in rice plants. This study suggested that the risk of accumulation of Ga and In in rice grains should be of concern when paddy rice is grown in acidic Ga- or In-contaminated soils with low Al availability.

Arf GAPs: A family of proteins with disparate functions that converge on a common structure, the integrin adhesion complex.

ADP-ribosylation factors (Arfs) are members of the Ras GTPase superfamily. The function of Arfs is dependent on GTPase-activating proteins (GAPs) and guanine nucleotide exchange factors (GEFs), which allow Arfs to cycle between the GDP-bound and GTP-bound forms. Arf GAPs have been shown to be present in integrin adhesion complexes, which include focal adhesions. Integrin adhesion complexes are composed of integrins, scaffolding proteins and signaling proteins and regulate cell proliferation, survival, differentiation and migration. Understanding the role of Arf GAPs in the regulation of integrin adhesion complexes is relevant to understanding normal physiology and cancer. In this review, we will discuss the contribution of the Arf GAP family members to the regulation of integrin adhesion complexes, examining the diverse mechanisms by which they control integrin adhesion complex formation, maturation and dissolution. GIT1 and ARAP2 serve as GAPs for Arf6, regulating Rac1 and other effectors by mechanisms still being defined. In contrast, GIT2 regulates Rac1 independent of Arf6. AGAP2 binds to and regulates focal adhesion kinase (FAK). ARAP2 and ACAP1, both Arf6 GAPs, regulate membrane trafficking of integrins through different endocytic pathways, exerting opposite effects on focal adhesions. ASAP1 not only regulates actin cytoskeleton remodeling through its interaction with nonmuscle myosin 2A, but is also important in integrin recycling. These examples illustrate the diversity and versatility of Arf GAPs as regulators of integrin adhesion complex structure and function.