scholarly journals Cluster of Differentiation Antigen 4 (CD4) Endocytosis and Adaptor Complex Binding Require Activation of the CD4 Endocytosis Signal by Serine Phosphorylation

1999 ◽  
Vol 10 (3) ◽  
pp. 677-691 ◽  
Author(s):  
Carol Pitcher ◽  
Stefan Höning ◽  
Anja Fingerhut ◽  
Katherine Bowers ◽  
Mark Marsh
Keyword(s):  
Phorbol Esters ◽  
Protein Complexes ◽  
Adaptor Protein ◽  
Cytoplasmic Domain ◽  
Serine Phosphorylation ◽  
Differentiation Antigen ◽  
Coated Pits ◽  
Early Endosomes ◽  
Clathrin Adaptor ◽  
Cluster Of Differentiation

Cluster of differentiation antigen 4 (CD4), the T lymphocyte antigen receptor component and human immunodeficiency virus coreceptor, is down-modulated when cells are activated by antigen or phorbol esters. During down-modulation CD4 dissociates from p56 lck , undergoes endocytosis through clathrin-coated pits, and is then sorted in early endosomes to late endocytic organelles where it is degraded. Previous studies have suggested that phosphorylation and a dileucine sequence are required for down-modulation. Using transfected HeLa cells, in which CD4 endocytosis can be studied in the absence of p56 lck , we show that the dileucine sequence in the cytoplasmic domain is essential for clathrin-mediated CD4 endocytosis. However, this sequence is only functional as an endocytosis signal when neighboring serine residues are phosphorylated. Phosphoserine is required for rapid endocytosis because CD4 molecules in which the cytoplasmic domain serine residues are substituted with glutamic acid residues are not internalized efficiently. Using surface plasmon resonance, we show that CD4 peptides containing the dileucine sequence bind weakly to clathrin adaptor protein complexes 2 and 1. The affinity of this interaction is increased 350- to 700-fold when the peptides also contain phosphoserine residues.

scholarly journals The alternate AP-1 adaptor subunit Apm2 interacts with the Mil1 regulatory protein and confers differential cargo sorting

2016 ◽  
Vol 27 (3) ◽  
pp. 588-598 ◽  
Author(s):  
Shawn T. Whitfield ◽  
Helen E. Burston ◽  
Björn D. M. Bean ◽  
Nandini Raghuram ◽  
Lymarie Maldonado-Báez ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Regulatory Protein ◽  
Adaptor Protein ◽  
General Mechanism ◽  
Specific Expression ◽  
Membrane Recruitment ◽  
Early Endosomes ◽  
Cell Type Specific Expression ◽  
Clathrin Adaptor ◽  
Cargo Sorting

Heterotetrameric adaptor protein complexes are important mediators of cargo protein sorting in clathrin-coated vesicles. The cell type–specific expression of alternate μ chains creates distinct forms of AP-1 with altered cargo sorting, but how these subunits confer differential function is unclear. Whereas some studies suggest the μ subunits specify localization to different cellular compartments, others find that the two forms of AP-1 are present in the same vesicle but recognize different cargo. Yeast have two forms of AP-1, which differ only in the μ chain. Here we show that the variant μ chain Apm2 confers distinct cargo-sorting functions. Loss of Apm2, but not of Apm1, increases cell surface levels of the v-SNARE Snc1. However, Apm2 is unable to replace Apm1 in sorting Chs3, which requires a dileucine motif recognized by the γ/σ subunits common to both complexes. Apm2 and Apm1 colocalize at Golgi/early endosomes, suggesting that they do not associate with distinct compartments. We identified a novel, conserved regulatory protein that is required for Apm2-dependent sorting events. Mil1 is a predicted lipase that binds Apm2 but not Apm1 and contributes to its membrane recruitment. Interactions with specific regulatory factors may provide a general mechanism to diversify the functional repertoire of clathrin adaptor complexes.

Clathrin, adaptors and eps15 in endosomes containing activated epidermal growth factor receptors

1999 ◽  
Vol 112 (3) ◽  
pp. 317-327 ◽  
Author(s):  
T. Sorkina ◽  
A. Bild ◽  
F. Tebar ◽  
A. Sorkin
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Adaptor Protein ◽  
Receptor Internalization ◽  
Image Deconvolution ◽  
Protein Markers ◽  
Coated Pits ◽  
Early Endosomes ◽  
Epidermal Growth ◽  
Clathrin Adaptor

Activation of the epidermal growth factor receptor (EGFR) by EGF results in binding of clathrin adaptor protein complex AP-2 to the receptor cytoplasmic tail. The transient interaction with AP-2 is thought to be responsible for the selective recruitment of the EGFR into coated pits during endocytosis. In this study we found that EGF-induced EGFR/AP-2 association, measured by co-immunoprecipitation, persists after receptor internalization. Double-label immunofluorescence of EGF-treated A-431 and COS-1 cells revealed the presence of AP-2, clathrin and eps15, another component of the plasma membrane coated pits, in the large perinuclear endosomes loaded with EGFRs. By optical sectioning and image deconvolution, the immunoreactivities were seen to be distributed within vesicular and tubular elements of these endosomes. In addition, these compartments contained the transferrin receptors and a EEA.1 protein, markers of early endosomes. Furthermore, Golgi clathrin adaptor complex AP-1 was found in EGFR-containing endosomes and EGFR immunoprecipitates in A-431 cells. The direct interaction of the EGFR with micro1 as well as micro2 subunits of AP-1 and AP-2, correspondingly, was shown using the yeast two-hybrid assay. Brefeldin A, a drug that releases AP-1 from the trans-Golgi membranes, had no effect on AP-1 association with endosomes and its co-precipitation with EGFR. Taken together, the data suggest that endosomal EGFR-AP complexes make up a significant portion of the total amount of these complexes detectable by co-immunoprecipitation. It can be proposed that APs are capable of binding to the endosomal membrane via a mechanism that requires AP interaction with the intracellular tails of multimeric receptors like activated EGFR, which in turn allows recruitment of clathrin and eps15. The hypothesis that the competition between adaptor complexes for binding to the receptor tails in endosomes may regulate of the sorting of receptors is discussed.

scholarly journals EHDS are serine phosphoproteins: EHD1 phosphorylation is enhanced by serum stimulation

2008 ◽  
Vol 13 (4) ◽  
Author(s):  
Boris Fichtman ◽  
Liat Ravid ◽  
Debora Rapaport ◽  
Mia Horowitz
Keyword(s):  
Adaptor Protein ◽  
Serine Phosphorylation ◽  
The Other ◽  
Serum Stimulation ◽  
Kinase C ◽  
Multiple Protein ◽  
Early Endosomes ◽  
Pkc Protein Kinase C ◽  
Clathrin Adaptor ◽  
Endocytic Pathways

AbstractEndocytic processes are mediated by multiple protein-protein interacting modules and regulated by phosphorylation and dephosphorylation. The Eps15 homology domain containing protein 1 (EHD1) has been implicated in regulating recycling of proteins, internalized both in clathrin-dependent and clathrin-independent endocytic pathways, from the recycling compartment to the plasma membrane. EHD1 was found in a complex with clathrin, adaptor protein complex-2 (AP-2) and insulin-like growth factor-1 receptor (IGF-1R), and was shown to interact with Rabenosyn-5, SNAP29, EHBP1 (EH domain binding protein 1) and syndapin I and II. In this study, we show that EHD1, like the other human EHDs, undergoes serine-phosphorylation. Our results also indicate that EHD1 is a serum-inducible serine-phosphoprotein and that PKC (protein kinase C) is one of its kinases. In addition, we show that inhibitors of clathrin-mediated endocytosis decrease EHD1 phosphorylation, while inhibitors of caveolinmediated endocytosis do not affect EHD1 phosphorylation. The results of experiments in which inhibitors of endocytosis were employed strongly suggest that EHD1 phosphorylation occurs between early endosomes and the endocytic recycling compartment.

scholarly journals Phosphoserine acidic cluster motifs bind distinct basic regions on the μ subunits of clathrin adaptor protein complexes

2018 ◽  
Vol 293 (40) ◽  
pp. 15678-15690 ◽  
Author(s):  
Rajendra Singh ◽  
Charlotte Stoneham ◽  
Christopher Lim ◽  
Xiaofei Jia ◽  
Javier Guenaga ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Adaptor Protein ◽  
Clathrin Adaptor

scholarly journals Regulation of Clathrin-Mediated Endocytosis

2018 ◽  
Vol 87 (1) ◽  
pp. 871-896 ◽  
Author(s):  
Marcel Mettlen ◽  
Ping-Hung Chen ◽  
Saipraveen Srinivasan ◽  
Gaudenz Danuser ◽  
Sandra L. Schmid
Keyword(s):  
Conformational Changes ◽  
Posttranslational Modifications ◽  
Mammalian Cells ◽  
Environmental Changes ◽  
Protein Complexes ◽  
Adaptor Protein ◽  
Endocytic Pathway ◽  
Coat Proteins ◽  
Membrane Composition ◽  
Coated Pits

Clathrin-mediated endocytosis (CME) is the major endocytic pathway in mammalian cells. It is responsible for the uptake of transmembrane receptors and transporters, for remodeling plasma membrane composition in response to environmental changes, and for regulating cell surface signaling. CME occurs via the assembly and maturation of clathrin-coated pits that concentrate cargo as they invaginate and pinch off to form clathrin-coated vesicles. In addition to the major coat proteins, clathrin triskelia and adaptor protein complexes, CME requires a myriad of endocytic accessory proteins and phosphatidylinositol lipids. CME is regulated at multiple steps—initiation, cargo selection, maturation, and fission—and is monitored by an endocytic checkpoint that induces disassembly of defective pits. Regulation occurs via posttranslational modifications, allosteric conformational changes, and isoform and splice-variant differences among components of the CME machinery, including the GTPase dynamin. This review summarizes recent findings on the regulation of CME and the evolution of this complex process.

scholarly journals Association and Colocalization of Eps15 with Adaptor Protein-2 and Clathrin

1997 ◽  
Vol 136 (4) ◽  
pp. 811-821 ◽  
Author(s):  
Sanne van Delft ◽  
Christopher Schumacher ◽  
Willem Hage ◽  
Arie J. Verkleij ◽  
Paul M.P. van Bergen en Henegouwen
Keyword(s):  
Egf Receptor ◽  
Critical Role ◽  
Adaptor Protein ◽  
Coated Vesicles ◽  
Endocytic Pathway ◽  
Egf Receptors ◽  
Coated Pits ◽  
Early Endosomes ◽  
Endosomal Membranes ◽  
Cell Fractions

Eps15 has been identified as a substrate of the EGF receptor tyrosine kinase. In this report, we show that activation of the EGF receptor by either EGF or TGF-α results in phosphorylation of Eps15. Stimulation of cells with PDGF or insulin did not lead to Eps15 phosphorylation, suggesting that phosphorylation of Eps15 is a receptor-specific process. We demonstrate that Eps15 is constitutively associated with both α-adaptin and clathrin. Upon EGF stimulation, Eps15 and α-adaptin are recruited to the EGF receptor. Using a truncated EGF receptor mutant, we demonstrate that the regulatory domain of the cytoplasmic tail of the EGF receptor is essential for the binding of Eps15. Fractionation studies reveal that Eps15 is present in cell fractions enriched for plasma membrane and endosomal membranes. Immunofluorescence studies show that Eps15 colocalizes with adaptor protein-2 (AP-2) and partially with clathrin. No colocalization of Eps15 was observed with the early endosomal markers rab4 and rab5. These observations indicate that Eps15 is present in coated pits and coated vesicles of the clathrin-mediated endocytic pathway, but not in early endosomes. Neither AP-2 nor clathrin are required for the binding of Eps15 to coated pits or coated vesicles, since in membranes lacking AP-2 and clathrin, Eps15 still shows the same staining pattern. These findings suggest that Eps15 may play a critical role in the recruitment of active EGF receptors into coated pit regions before endocytosis of ligand-occupied EGF receptors.

scholarly journals Two Clathrin Adaptor Protein Complexes Instruct Axon-Dendrite Polarity

Neuron ◽  
2016 ◽  
Vol 90 (3) ◽  
pp. 564-580 ◽  
Author(s):  
Pengpeng Li ◽  
Sean A. Merrill ◽  
Erik M. Jorgensen ◽  
Kang Shen
Keyword(s):  
Protein Complexes ◽  
Adaptor Protein ◽  
Clathrin Adaptor

scholarly journals Functions of Adaptor Protein (AP)-3 and AP-1 in Tyrosinase Sorting from Endosomes to Melanosomes

2005 ◽  
Vol 16 (11) ◽  
pp. 5356-5372 ◽  
Author(s):  
Alexander C. Theos ◽  
Danièle Tenza ◽  
José A. Martina ◽  
Ilse Hurbain ◽  
Andrew A. Peden ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Adaptor Protein ◽  
Specific Protein ◽  
Model System ◽  
Multivesicular Bodies ◽  
Substantial Fraction ◽  
Sorting Signal ◽  
Early Endosomes ◽  
Unique Model ◽  
Multivesicular Endosomes

Specialized cells exploit adaptor protein complexes for unique post-Golgi sorting events, providing a unique model system to specify adaptor function. Here, we show that AP-3 and AP-1 function independently in sorting of the melanocyte-specific protein tyrosinase from endosomes to the melanosome, a specialized lysosome-related organelle distinguishable from lysosomes. AP-3 and AP-1 localize in melanocytes primarily to clathrin-coated buds on tubular early endosomes near melanosomes. Both adaptors recognize the tyrosinase dileucine-based melanosome sorting signal, and tyrosinase largely colocalizes with each adaptor on endosomes. In AP-3-deficient melanocytes, tyrosinase accumulates inappropriately in vacuolar and multivesicular endosomes. Nevertheless, a substantial fraction still accumulates on melanosomes, concomitant with increased association with endosomal AP-1. Our data indicate that AP-3 and AP-1 function in partially redundant pathways to transfer tyrosinase from distinct endosomal subdomains to melanosomes and that the AP-3 pathway ensures that tyrosinase averts entrapment on internal membranes of forming multivesicular bodies.

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