"In vitro, ACAP1 and ACAP2 preferred Arf6 as a substrate, rather than Arf1 and Arf5, more so than did ASAP1."

"[R448Q]ACAP1 and [R442Q]ACAP2 had no detectable activity using Arf6 as a substrate"

"ACAP1 and ACAP2 are preferentially activated by phosphatidylinositol 4,5-bisphosphate [PIP2; not shown] and do contain GAP activity."

"In untreated cells, although ACAP1 and ACAP2 were primarily cytosolic, they could be observed on tubular portions of the Arf6 endosomal compartment"

"ACAP1 remained cytosolic and was not recruited to the endosome or the plasma membrane in cells expressing the dominant-negative Arf6, T27N, even in the presence of AlF4"

"During PDGF stimulation of NIH 3T3 cells, ACAP1, ACAP2, and ASAP1 colocalized in dorsal ruffles and at the edge of resting and protrusive HeLa cells."

"A point mutant of ACAP1 that lacked GAP activity was not observed on the tubular endosomes, but rather was associated with the plasma membrane protrusions"

"In HeLa cells, overexpression of either ACAP blocked the formation of Arf6-dependent protrusions."

"We now show that ACAP1 also participates in the regulated recycling of integrin beta1 to control cell migration."

"the role of ACAP1 in beta1 recycling requires its phosphorylation by Akt"

"Disrupting the activities of either ACAP1 or Akt, or their assembly with endosomal beta1, inhibits beta1 recycling and cell migration."

"Second, GULP associated with the Arf6-GAP ACAP1 at endogenous levels."

"Third, GULP reversed the Arf6-GTP decrease induced by ACAP1, and countered the ACAP1-mediated inhibition of cell migration."

"Fourth, GULP, ACAP1, and GDP-bound Arf6 were part of a tripartite complex, suggesting sequestration of ACAP1 as one mechanism of GULP action."

"The Arf6-specific GAP ACAP1 is a negative regulator of Arf6-mediated signaling"

"We find that ACAP1, a GTPase-activating protein (GAP) for ADP-ribosylation factor (ARF) 6, is part of a novel clathrin coat complex that is regulated by ARF6 for endocytic recycling in two key physiological settings, stimulation-dependent recycling of integrin that is critical for cell migration and insulin-stimulated recycling of glucose transporter type 4 (Glut4), which is required for glucose homeostasis."

"We initially defined a critical sequence in the cytoplasmic domain of integrin β1 recognized by ACAP1 and showed that this sequence acts as a recycling sorting signal."

"ACAP1 (ArfGAP with coiled-coil, ankyrin repeat, and PH domains protein 1) is an ARF6 GAP that also acts as a key component of a recently defined clathrin complex for endocytic recycling."

"which suggest that phosphorylation of ACAP1 relieves a localized mechanism of autoinhibition in regulating cargo binding"

"Here, we show that this BAR domain can neither bind membrane nor impart curvature, but instead requires a neighboring PH (Pleckstrin Homology) domain to achieve these functions."

"Specific residues within the PH domain are responsible for both membrane binding and curvature generation."

"The BAR domain adjacent to the PH domain instead interacts with the BAR domains of neighboring ACAP1 proteins to enable clustering at the membrane."

"the BAR domain of ACAP1 also showed little affinity to the generated liposomes, regardless of their size"

"We next found that ACAP1BAR-PH also induces membrane curvature, as reflected by the tubulation of liposomes, which was visualized by negative-stain electron microscopy (EM)"

"In particular, mutation of this residue to alanine (F280A) reduced the ability of ACAP1BAR-PH to induce liposome tubulation, while a more conservative mutation (F280W) preserved the ability to tubulate liposomes."

"Second, there is no significant interaction between the BAR domain and the underlying membrane."

"Instead, the BAR domain contributes to the packing interfaces among ACAP1BAR-PH protein in assembling the coating on liposome membrane."

"Dimerization of ACAP1BAR-PH was confirmed from size exclusion chromatography and light scattering"

"The in vitro disassociation constant of the dimerization was calculated to be below 5 µM, according to the quantitative gel-filtration experiment"

"The dimer interface has a large area (4100 Å2), and contains mainly hydrophobic residues that likely mediate dimerization."

"this additional finding suggests that one PH domain is sufficient to confer the ability of the ACAP1BAR-PH dimer to insert into membrane and impart curvature"

"ARAP2 formed a complex with APPL1 and colocalized with Arf6 and APPL in a compartment distinct from the Arf6/ACAP1 tubular recycling endosome."

"ARAP2 knockdown slowed, whereas ACAP1 knockdown accelerated, integrin β1 internalization."

"ARAP2 overexpression promoted large FAs, but ACAP1 overexpression reduced FAs."

"While there were few tubules induced by the expression of ACAP1 alone, numerous endosomal tubules were induced by coexpression of PIP5K and ACAP1."

"Truncated and point mutations in the ACAP1 BAR and PH domains revealed that both BAR and PH domains are required for tubulation."

"A group of 12 Src homology (SH) 3 domains from eight human proteins (Swiss-Prot ID: SRC, PLCG1, P85A, NCK1, GRB2, FYN, CRK) were used to screen a peptide target array composed of 1536 potential ligands, which led to the identification of 921 binary interactions between these proteins and 284 targets."

"We extended an established yeast two-hybrid system employing human protein kinases for the analyses of phospho-tyrosine (pY)-dependent PPIs in a direct experimental, large-scale approach."

"We identified 292 mostly novel pY-dependent PPIs"

"the majority of PPIs in HuRI were found in only one screen"

"While HuRI displays highly significant overlap with known functional relationships, the cellular function of most individual PPIs remains to be elucidated."

"connects ∼5,000 human proteins via ∼30,000 candidate interactions and is generated by systematic yeast two-hybrid interaction screening of ∼500 ND-related proteins and integration of literature interactions"

"Isolated membranes were treated with reagents that have been reported to remove peripheral membrane proteins."

"The remaining species were largely involved in cellular processes and molecular functions that could be predicted to be transiently associated with membranes."