scholarly journals The endosomal adaptor protein APPL1 impairs the turnover of leading edge adhesions to regulate cell migration

2012 ◽  
Vol 23 (8) ◽  
pp. 1486-1499 ◽  
Author(s):  
Joshua A. Broussard ◽  
Wan-hsin Lin ◽  
Devi Majumdar ◽  
Bridget Anderson ◽  
Brady Eason ◽  
...  
Keyword(s):  
Cell Migration ◽  
Leucine Zipper ◽  
Adaptor Protein ◽  
Leading Edge ◽  
Adaptor Proteins ◽  
Ph Domain ◽  
Serine Threonine Kinase ◽  
Nonreceptor Tyrosine Kinase ◽  
Complex Process ◽  
Flow Of Information

Cell migration is a complex process that requires the integration of signaling events that occur in distinct locations within the cell. Adaptor proteins, which can localize to different subcellular compartments, where they bring together key signaling proteins, are emerging as attractive candidates for controlling spatially coordinated processes. However, their function in regulating cell migration is not well understood. In this study, we demonstrate a novel role for the adaptor protein containing a pleckstrin-homology (PH) domain, phosphotyrosine-binding (PTB) domain, and leucine zipper motif 1 (APPL1) in regulating cell migration. APPL1 impairs migration by hindering the turnover of adhesions at the leading edge of cells. The mechanism by which APPL1 regulates migration and adhesion dynamics is by inhibiting the activity of the serine/threonine kinase Akt at the cell edge and within adhesions. In addition, APPL1 significantly decreases the tyrosine phosphorylation of Akt by the nonreceptor tyrosine kinase Src, which is critical for Akt-mediated cell migration. Thus, our results demonstrate an important new function for APPL1 in regulating cell migration and adhesion turnover through a mechanism that depends on Src and Akt. Moreover, our data further underscore the importance of adaptor proteins in modulating the flow of information through signaling pathways.

scholarly journals Crk adaptor proteins mediate actin-dependent T cell migration and mechanosensing induced by the integrin LFA-1

2018 ◽  
Vol 11 (560) ◽  
pp. eaat3178 ◽  
Author(s):  
Nathan H. Roy ◽  
Joanna L. MacKay ◽  
Tanner F. Robertson ◽  
Daniel A. Hammer ◽  
Janis K. Burkhardt
Keyword(s):  
T Cells ◽  
Cell Migration ◽  
T Cell ◽  
Actin Polymerization ◽  
Adaptor Protein ◽  
Leading Edge ◽  
Adaptor Proteins ◽  
Substrate Stiffness ◽  
Lymphocyte Function ◽  
Molecular Events

T cell entry into inflamed tissue involves firm adhesion, spreading, and migration of the T cells across endothelial barriers. These events depend on “outside-in” signals through which engaged integrins direct cytoskeletal reorganization. We investigated the molecular events that mediate this process and found that T cells from mice lacking expression of the adaptor protein Crk exhibited defects in phenotypes induced by the integrin lymphocyte function–associated antigen 1 (LFA-1), namely, actin polymerization, leading edge formation, and two-dimensional cell migration. Crk protein was an essential mediator of LFA-1 signaling–induced phosphorylation of the E3 ubiquitin ligase c-Cbl and its subsequent interaction with the phosphatidylinositol 3-kinase (PI3K) subunit p85, thus promoting PI3K activity and cytoskeletal remodeling. In addition, we found that Crk proteins were required for T cells to respond to changes in substrate stiffness, as measured by alterations in cell spreading and differential phosphorylation of the force-sensitive protein CasL. These findings identify Crk proteins as key intermediates coupling LFA-1 signals to actin remodeling and provide mechanistic insights into how T cells sense and respond to substrate stiffness.

scholarly journals APPL1 is a multifunctional endosomal signaling adaptor protein

2017 ◽  
Vol 45 (3) ◽  
pp. 771-779 ◽  
Author(s):  
Nicole L. Diggins ◽  
Donna J. Webb
Keyword(s):  
Signaling Pathways ◽  
Protein Interactions ◽  
Leucine Zipper ◽  
Adaptor Protein ◽  
Adaptor Proteins ◽  
Ph Domain ◽  
Protein Protein Interactions ◽  
Multiple Protein ◽  
Signaling Receptors ◽  
Ptb Domain

Endosomal adaptor proteins are important regulators of signaling pathways underlying many biological processes. These adaptors can integrate signals from multiple pathways via localization to specific endosomal compartments, as well as through multiple protein–protein interactions. One such adaptor protein that has been implicated in regulating signaling pathways is the adaptor protein containing a pleckstrin homology (PH) domain, phosphotyrosine-binding (PTB) domain, and leucine zipper motif 1 (APPL1). APPL1 localizes to a subset of Rab5-positive endosomes through its Bin–Amphiphysin–Rvs and PH domains, and it coordinates signaling pathways through its interaction with many signaling receptors and proteins through its PTB domain. This review discusses our current understanding of the role of APPL1 in signaling and trafficking, as well as highlights recent work into the function of APPL1 in cell migration and adhesion.

scholarly journals Rapid whole cell imaging reveals a calcium-APPL1-dynein nexus that regulates cohort trafficking of stimulated EGF receptors

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
H. M. York ◽  
A. Patil ◽  
U. K. Moorthi ◽  
A. Kaur ◽  
A. Bhowmik ◽  
...  
Keyword(s):  
Signal Processing ◽  
Growth Factor ◽  
Leucine Zipper ◽  
Adaptor Protein ◽  
Endosomal Trafficking ◽  
Egf Receptors ◽  
Ph Domain ◽  
Cell Level ◽  
Whole Cell ◽  
Light Sheet Microscopy

AbstractThe endosomal system provides rich signal processing capabilities for responses elicited by growth factor receptors and their ligands. At the single cell level, endosomal trafficking becomes a critical component of signal processing, as exemplified by the epidermal growth factor (EGF) receptors. Activated EGFRs are trafficked to the phosphatase-enriched peri-nuclear region (PNR), where they are dephosphorylated and degraded. The details of the mechanisms that govern the movements of stimulated EGFRs towards the PNR, are not completely known. Here, exploiting the advantages of lattice light-sheet microscopy, we show that EGFR activation by EGF triggers a transient calcium increase causing a whole-cell level redistribution of Adaptor Protein, Phosphotyrosine Interacting with PH Domain And Leucine Zipper 1 (APPL1) from pre-existing endosomes within one minute, the rebinding of liberated APPL1 directly to EGFR, and the dynein-dependent translocation of APPL1-EGF-bearing endosomes to the PNR within ten minutes. The cell spanning, fast acting network that we reveal integrates a cascade of events dedicated to the cohort movement of activated EGF receptors. Our findings support the intriguing proposal that certain endosomal pathways have shed some of the stochastic strategies of traditional trafficking and have evolved processes that provide the temporal predictability that typify canonical signaling.

scholarly journals Filopodia formation and endosome clustering induced by mutant plus-end–directed myosin VI

2017 ◽  
Vol 114 (7) ◽  
pp. 1595-1600 ◽  
Author(s):  
Thomas A. Masters ◽  
Folma Buss
Keyword(s):  
Actin Filaments ◽  
Leucine Zipper ◽  
Adaptor Protein ◽  
Cell Cortex ◽  
Myosin Vi ◽  
Actin Network ◽  
Ph Domain ◽  
Cortical Actin ◽  
Directed Movement ◽  
Interacting Protein

Myosin VI (MYO6) is the only myosin known to move toward the minus end of actin filaments. It has roles in numerous cellular processes, including maintenance of stereocilia structure, endocytosis, and autophagosome maturation. However, the functional necessity of minus-end–directed movement along actin is unclear as the underlying architecture of the local actin network is often unknown. To address this question, we engineered a mutant of MYO6, MYO6+, which undergoes plus-end–directed movement while retaining physiological cargo interactions in the tail. Expression of this mutant motor in HeLa cells led to a dramatic reorganization of cortical actin filaments and the formation of actin-rich filopodia. MYO6 is present on peripheral adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper 1 (APPL1) signaling endosomes and MYO6+ expression causes a dramatic relocalization and clustering of this endocytic compartment in the cell cortex. MYO6+ and its adaptor GAIP interacting protein, C terminus (GIPC) accumulate at the tips of these filopodia, while APPL1 endosomes accumulate at the base. A combination of MYO6+ mutagenesis and siRNA-mediated depletion of MYO6 binding partners demonstrates that motor activity and binding to endosomal membranes mediated by GIPC and PI(4,5)P2 are crucial for filopodia formation. A similar reorganization of actin is induced by a constitutive dimer of MYO6+, indicating that multimerization of MYO6 on endosomes through binding to GIPC is required for this cellular activity and regulation of actin network structure. This unique engineered MYO6+ offers insights into both filopodia formation and MYO6 motor function at endosomes and at the plasma membrane.

scholarly journals Deficiency in the endocytic adaptor protein PHETA1/2 impairs renal and craniofacial development

2019 ◽  
Author(s):  
Kristin M. Ates ◽  
Tong Wang ◽  
Trevor Moreland ◽  
Rajalakshmi Veeranan-Karmegam ◽  
Priya Anand ◽  
...  
Keyword(s):  
De Novo ◽  
Adaptor Protein ◽  
Adaptor Proteins ◽  
Craniofacial Development ◽  
Dominant Negative ◽  
Patient Specific ◽  
Ph Domain ◽  
Causative Gene ◽  
Chondrocyte Maturation

AbstractA critical barrier in the treatment of endosomal and lysosomal diseases is the lack of understanding of the in vivo functions of the putative causative genes. We addressed this by investigating a key pair of endocytic adaptor proteins, PH domain containing endocytic trafficking adaptor 1 and 2 (PHETA1/2, also known as FAM109A/B, Ses1/2, IPIP27A/B), which interact with the protein product of OCRL, the causative gene for Lowe syndrome. Here we conducted the first study of PHETA1/2 in vivo, utilizing the zebrafish system. We found that impairment of both zebrafish orthologs, pheta1 and pheta2, disrupted endocytosis and ciliogenesis. In addition, pheta1/2 mutant animals exhibited reduced jaw size and delayed chondrocyte maturation, indicating a role in craniofacial development. Deficiency of pheta1/2 resulted in dysregulation of cathepsin K, which led to an increased abundance of type II collagen in craniofacial cartilages. The abnormal renal and craniofacial phenotypes in the pheta1/2 mutant animals were consistent with the clinical presentations of a patient with a de novo arginine (R) to cysteine (C) variant (R6C) of PHETA1. Expressing the patient-specific variant in zebrafish exacerbated craniofacial deficits, suggesting that the R6C allele acts in a dominant-negative manner. Together, these results provide insights into the in vivo roles of PHETA1/2 and suggest that the R6C variant is contributory to the pathogenesis of disease in the patient.

scholarly journals Role of Phosphatidylinositol 3′ Kinase and a Downstream Pleckstrin Homology Domain–Containing Protein in Controlling Chemotaxis inDictyostelium

2001 ◽  
Vol 153 (4) ◽  
pp. 795-810 ◽  
Author(s):  
Satoru Funamoto ◽  
Kristina Milan ◽  
Ruedi Meili ◽  
Richard A. Firtel
Keyword(s):  
Actin Polymerization ◽  
Adaptor Protein ◽  
Leading Edge ◽  
Membrane Lipid ◽  
Lipid Binding ◽  
Pleckstrin Homology Domain ◽  
Ph Domain ◽  
Wild Type ◽  
Pi3k Inhibitor Ly294002 ◽  
Effector Pathways

We show that cells lacking two Dictyostelium class I phosphatidylinositol (PI) 3′ kinases (PI3K and pi3k1/2-null cells) or wild-type cells treated with the PI3K inhibitor LY294002 are unable to properly polarize, are very defective in the temporal, spatial, and quantitative regulation of chemoattractant-mediated filamentous (F)-actin polymerization, and chemotax very slowly. PI3K is thought to produce membrane lipid-binding sites for localization of PH domain–containing proteins. We demonstrate that in response to chemoattractants three PH domain–containing proteins do not localize to the leading edge in pi3k1/2-null cells, and the translocation is blocked in wild-type cells by LY294002. Cells lacking one of these proteins, phdA-null cells, exhibit defects in the level and kinetics of actin polymerization at the leading edge and have chemotaxis phenotypes that are distinct from those described previously for protein kinase B (PKB) (pkbA)-null cells. Phenotypes of PhdA-dominant interfering mutations suggest that PhdA is an adaptor protein that regulates F-actin localization in response to chemoattractants and links PI3K to the control of F-actin polymerization at the leading edge during pseudopod formation. We suggest that PKB and PhdA lie downstream from PI3K and control different downstream effector pathways that are essential for proper chemotaxis.

Adaptor protein containing PH domain, PTB domain and leucine zipper (APPL1) regulates the protein level of EGFR by modulating its trafficking

2011 ◽  
Vol 415 (1) ◽  
pp. 206-211 ◽  
Author(s):  
Jae-Rin Lee ◽  
Hwa-Sun Hahn ◽  
Young-Hoon Kim ◽  
Hong-Hoa Nguyen ◽  
Jun-Mo Yang ◽  
...  
Keyword(s):  
Protein Level ◽  
Leucine Zipper ◽  
Adaptor Protein ◽  
Ph Domain ◽  
Ptb Domain

scholarly journals SUN-687 MODY-14: A Rare Cause of Inherited Diabetes

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Riley Epp ◽  
Amel Arnaout ◽  
Mary-Anne Doyle
Keyword(s):  
Insulin Secretion ◽  
Leucine Zipper ◽  
Novel Mutation ◽  
Adaptor Protein ◽  
Alpha Helix ◽  
Ph Domain ◽  
Loss Of Function ◽  
Physically Active ◽  
Bar Domain ◽  
Oral Hypoglycemic Medications

Abstract Case: A 29-year-old physically active, lean man presented with a three-year history of poorly controlled diabetes diagnosed during a flu-like illness and no history diabetic ketoacidosis. Despite treatment with various oral hypoglycemic medications his glycated hemoglobin was consistently above target (never falling below 9.3%) and correlated with home blood glucose readings. His mother had gestational diabetes with all three of her pregnancies. His maternal grandmother was also known to have diabetes. Pancreatic islet antibodies were negative. C-peptide levels were low. Monogenic diabetes (MODY) was suspected. Discussion: Genetic testing revealed a mutation in the Adaptor Protein, Phosphotyrosine Interaction, PH domain, and leucine zipper containing 1 (APPL1) gene. APPL1 is involved in insulin secretion and insulin signaling via the Akt pathway [1,2,3]. Other mutations of APPL1 causing MODY-14 have been previously described [4]. In our case, the substitution of glutamate for aspartate at the 265 position occurs within the unique fourth alpha helix of the BAR domain of APPL1 [5]. Generally, BAR domains contribute to APPL1 dimerization and plasma membrane association [5]. Substitution of glutamate could disrupt protein folding, impairing dimerization and phosphorylation of Akt, thereby decreased insulin secretion as seen in other mutations of the BAR domain of APPL1 [4]. Although this remains a hypothesis, loss-of-function of APPL1 would explain the lack of insulin secretion without evidence of pancreatic autoimmunity. If the hypothesis is correct, it would be a novel mutation attributable to this MODY subtype. 1. Cheng KKY, Lam KSL, Wu D, et al. APPL1 potentiates insulin secretion in pancreatic β cells by enhancing protein kinase Akt-dependent expression of SNARE proteins in mice. Proc Natl Acad Sci U S A. 2012;109(23):8919–8924. doi:10.1073/pnas.1202435109 2. Ryu J, Galan AK, Xin X, et al. APPL1 potentiates insulin sensitivity by facilitating the binding of IRS1/2 to the insulin receptor. Cell Rep. 2014;7(4):1227–1238. doi:10.1016/j.celrep.2014.04.006 3. Saito T, Jones CC, Huang S, Czech MP, Pilch PF. The interaction of Akt with APPL1 is required for insulin-stimulated Glut4 translocation. J Biol Chem. 2007;282(44):32280–32287. doi:10.1074/jbc.M704150200 4. Prudente S, Jungtrakoon P, Marucci A, et al. Loss-of-Function Mutations in APPL1 in Familial Diabetes Mellitus. Am J Hum Genet. 2015;97(1):177–185. doi:10.1016/j.ajhg.2015.05.011 5. Li J, Mao X, Dong LQ, Liu F, Tong L. Crystal Structures of the BAR-PH and PTB Domains of Human APPL1. Structure. 2007;15(5):525–533. doi:10.1016/j.str.2007.03.011

scholarly journals The Arf6 GTPase-activating Proteins ARAP2 and ACAP1 Define Distinct Endosomal Compartments That Regulate Integrin α5β1 Traffic

2014 ◽  
Vol 289 (44) ◽  
pp. 30237-30248 ◽  
Author(s):  
Pei-Wen Chen ◽  
Ruibai Luo ◽  
Xiaoying Jian ◽  
Paul A. Randazzo
Keyword(s):  
Leucine Zipper ◽  
Focal Adhesions ◽  
Adaptor Protein ◽  
Ph Domain ◽  
Integrin Β1 ◽  
Recycling Endosome ◽  
Gtpase Activating Proteins ◽  
Cell Adhesion And Migration ◽  
And Migration ◽  
Opposing Effects

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

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