Antibody data
- Antibody Data
- Antigen structure
- References [92]
- Comments [0]
- Validations
- Western blot [3]
- Immunocytochemistry [3]
- Immunohistochemistry [2]
- Other assay [17]
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Validation data
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- Product number
- MA1-065 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Clathrin Heavy Chain Monoclonal Antibody (X22)
- Antibody type
- Monoclonal
- Antigen
- Purifed from natural sources
- Description
- MA1-065 detects clathrin heavy chain in non-human primate, bovine, human, rat and mouse tissues as well as hamster (CHO) cells.
- Antibody clone number
- X22
- Concentration
- 6 mg/mL
Submitted references The obligate intracellular bacterium Orientia tsutsugamushi differentiates into a developmentally distinct extracellular state.
Visualizing cellular and tissue ultrastructure using Ten-fold Robust Expansion Microscopy (TREx).
Vibropolyfection: coupling polymer-mediated gene delivery to mechanical stimulation to enhance transfection of adherent cells.
The nanoscale molecular morphology of docked exocytic dense-core vesicles in neuroendocrine cells.
TGFβ receptor endocytosis and Smad signaling require synaptojanin1, PI3K-C2α-, and INPP4B-mediated phosphoinositide conversions.
Physiological blood-brain transport is impaired with age by a shift in transcytosis.
HHV-7 U21 exploits Golgi quality control carriers to reroute class I MHC molecules to lysosomes.
Structurally distinct endocytic pathways for B cell receptors in B lymphocytes.
Acute Statin Treatment Improves Antibody Accumulation in EGFR- and PSMA-Expressing Tumors.
A unique role for clathrin light chain A in cell spreading and migration.
Chlamydia exploits filopodial capture and a macropinocytosis-like pathway for host cell entry.
Rare Disease Mechanisms Identified by Genealogical Proteomics of Copper Homeostasis Mutant Pedigrees.
A Flat BAR Protein Promotes Actin Polymerization at the Base of Clathrin-Coated Pits.
ELMOD1 Stimulates ARF6-GTP Hydrolysis to Stabilize Apical Structures in Developing Vestibular Hair Cells.
An Aberrant Phosphorylation of Amyloid Precursor Protein Tyrosine Regulates Its Trafficking and the Binding to the Clathrin Endocytic Complex in Neural Stem Cells of Alzheimer's Disease Patients.
Surface Toll-like receptor 3 expression in metastatic intestinal epithelial cells induces inflammatory cytokine production and promotes invasiveness.
Influenza virus genome reaches the plasma membrane via a modified endoplasmic reticulum and Rab11-dependent vesicles.
Flat clathrin lattices are dynamic actin-controlled hubs for clathrin-mediated endocytosis and signalling of specific receptors.
Nanoscale manipulation of membrane curvature for probing endocytosis in live cells.
Endocytic Pathways Used by Andes Virus to Enter Primary Human Lung Endothelial Cells.
madSTORM: a superresolution technique for large-scale multiplexing at single-molecule accuracy.
Caveolin- and clathrin-independent entry of BKPyV into primary human proximal tubule epithelial cells.
Synaptotagmin-11 inhibits clathrin-mediated and bulk endocytosis.
High affinity receptor labeling based on basic leucine zipper domain peptides conjugated with pH-sensitive fluorescent dye: Visualization of AMPA-type glutamate receptor endocytosis in living neurons.
Diverse functions of myosin VI elucidated by an isoform-specific α-helix domain.
Axial superresolution via multiangle TIRF microscopy with sequential imaging and photobleaching.
Central role for PICALM in amyloid-β blood-brain barrier transcytosis and clearance.
Endophilin marks and controls a clathrin-independent endocytic pathway.
Lack of CD2AP disrupts Glut4 trafficking and attenuates glucose uptake in podocytes.
Uptake of Helicobacter pylori vesicles is facilitated by clathrin-dependent and clathrin-independent endocytic pathways.
Characterization of human astrovirus cell entry.
The X-linked mental retardation protein OPHN1 interacts with Homer1b/c to control spine endocytic zone positioning and expression of synaptic potentiation.
Functional coupling of chloride-proton exchanger ClC-5 to gastric H+,K+-ATPase.
Alternative endocytosis pathway for productive entry of hepatitis C virus.
Stabilization of actin bundles by a dynamin 1/cortactin ring complex is necessary for growth cone filopodia.
Netrin-1-induced activation of Notch signaling mediates glioblastoma cell invasion.
The LXR-IDOL axis defines a clathrin-, caveolae-, and dynamin-independent endocytic route for LDLR internalization and lysosomal degradation.
The PDZ protein GIPC regulates trafficking of the LPA1 receptor from APPL signaling endosomes and attenuates the cell's response to LPA.
Cell surface ceramide controls translocation of transferrin receptor to clathrin-coated pits.
Synaptotagmin 1 is necessary for the Ca2+ dependence of clathrin-mediated endocytosis.
Adaptor protein 2-mediated endocytosis of the β-secretase BACE1 is dispensable for amyloid precursor protein processing.
Rift Valley fever virus strain MP-12 enters mammalian host cells via caveola-mediated endocytosis.
Multi-colour direct STORM with red emitting carbocyanines.
Sphingomyelin synthase 1-generated sphingomyelin plays an important role in transferrin trafficking and cell proliferation.
Rapid and efficient clathrin-mediated endocytosis revealed in genome-edited mammalian cells.
Fast, three-dimensional super-resolution imaging of live cells.
Intracellular trafficking of hyaluronic acid-chitosan oligomer-based nanoparticles in cultured human ocular surface cells.
Role of the clathrin terminal domain in regulating coated pit dynamics revealed by small molecule inhibition.
Coupling between clathrin-dependent endocytic budding and F-BAR-dependent tubulation in a cell-free system.
Ebola virus uses clathrin-mediated endocytosis as an entry pathway.
EphA8-ephrinA5 signaling and clathrin-mediated endocytosis is regulated by Tiam-1, a Rac-specific guanine nucleotide exchange factor.
Ornithine decarboxylase antizyme inhibitor 2 regulates intracellular vesicle trafficking.
Disruption of zebrafish cyclin G-associated kinase (GAK) function impairs the expression of Notch-dependent genes during neurogenesis and causes defects in neuronal development.
Assembly of connexin43 into gap junctions is regulated differentially by E-cadherin and N-cadherin in rat liver epithelial cells.
Rapid turnover of spinules at synaptic terminals.
Phosphoinositide 3-kinase p110beta activity: key role in metabolism and mammary gland cancer but not development.
Hrs and SNX3 functions in sorting and membrane invagination within multivesicular bodies.
Mass spectrometry analysis of the native protein complex containing actinin-4 in prostate cancer cells.
A role for clathrin in reassembly of the Golgi apparatus.
Role of internalization of M2 muscarinic receptor via clathrin-coated vesicles in desensitization of the muscarinic K+ current in heart.
GBF1, a cis-Golgi and VTCs-localized ARF-GEF, is implicated in ER-to-Golgi protein traffic.
Interaction of SPIN90 with syndapin is implicated in clathrin-mediated endocytic pathway in fibroblasts.
Trimerisation is important for the function of clathrin at the mitotic spindle.
Epidermal growth factor receptor exposed to oxidative stress undergoes Src- and caveolin-1-dependent perinuclear trafficking.
ST6Gal-I restrains CD22-dependent antigen receptor endocytosis and Shp-1 recruitment in normal and pathogenic immune signaling.
Vascular endothelial cadherin controls VEGFR-2 internalization and signaling from intracellular compartments.
SPG3A protein atlastin-1 is enriched in growth cones and promotes axon elongation during neuronal development.
Exchange of clathrin, AP2 and epsin on clathrin-coated pits in permeabilized tissue culture cells.
Analysis of foot-and-mouth disease virus internalization events in cultured cells.
Clathrin-independent endocytosis of ubiquitinated cargos.
TTP specifically regulates the internalization of the transferrin receptor.
A novel GTPase-activating protein for ARF6 directly interacts with clathrin and regulates clathrin-dependent endocytosis.
Increased expression of protein C-mannosylation in the aortic vessels of diabetic Zucker rats.
Actin dynamics coupled to clathrin-coated vesicle formation at the trans-Golgi network.
Endocytosis of epithelial apical junctional proteins by a clathrin-mediated pathway into a unique storage compartment.
Differences in endosomal targeting of human (beta)1- and (beta)2-adrenergic receptors following clathrin-mediated endocytosis.
Detergent-resistant membrane microdomains facilitate Ib oligomer formation and biological activity of Clostridium perfringens iota-toxin.
RNAi-mediated Hip1R silencing results in stable association between the endocytic machinery and the actin assembly machinery.
Targeted chemical disruption of clathrin function in living cells.
PP1 inhibitor induces degradation of RETMEN2A and RETMEN2B oncoproteins through proteosomal targeting.
Spatial regulation of Galphai protein signaling in clathrin-coated membrane microdomains containing GAIP.
Sustained stimulation shifts the mechanism of endocytosis from dynamin-1-dependent rapid endocytosis to clathrin- and dynamin-2-mediated slow endocytosis in chromaffin cells.
HIP1 functions in clathrin-mediated endocytosis through binding to clathrin and adaptor protein 2.
Dynamin is involved in human epithelial cell vacuolation caused by the Helicobacter pylori-produced cytotoxin VacA.
Hsc70 chaperones clathrin and primes it to interact with vesicle membranes.
Role of cyclin G-associated kinase in uncoating clathrin-coated vesicles from non-neuronal cells.
Clathrin-coated vesicles bearing GAIP possess GTPase-activating protein activity in vitro.
Overexpression of proteins containing tyrosine- or leucine-based sorting signals affects transferrin receptor trafficking.
An actin-binding protein of the Sla2/Huntingtin interacting protein 1 family is a novel component of clathrin-coated pits and vesicles.
Characterization of the adaptor-related protein complex, AP-3.
Targeting and mistargeting of plasma membrane adaptors in vitro.
100-kD proteins of Golgi- and trans-Golgi network-associated coated vesicles have related but distinct membrane binding properties.
Atwal S, Wongsantichon J, Giengkam S, Saharat K, Pittayasathornthun YJ, Chuenklin S, Wang LC, Chung T, Huh H, Lee SH, Sobota RM, Salje J
Nature communications 2022 Jun 23;13(1):3603
Nature communications 2022 Jun 23;13(1):3603
Visualizing cellular and tissue ultrastructure using Ten-fold Robust Expansion Microscopy (TREx).
Damstra HGJ, Mohar B, Eddison M, Akhmanova A, Kapitein LC, Tillberg PW
eLife 2022 Feb 18;11
eLife 2022 Feb 18;11
Vibropolyfection: coupling polymer-mediated gene delivery to mechanical stimulation to enhance transfection of adherent cells.
Ponti F, Bono N, Russo L, Bigini P, Mantovani D, Candiani G
Journal of nanobiotechnology 2022 Aug 6;20(1):363
Journal of nanobiotechnology 2022 Aug 6;20(1):363
The nanoscale molecular morphology of docked exocytic dense-core vesicles in neuroendocrine cells.
Prasai B, Haber GJ, Strub MP, Ahn R, Ciemniecki JA, Sochacki KA, Taraska JW
Nature communications 2021 Jun 25;12(1):3970
Nature communications 2021 Jun 25;12(1):3970
TGFβ receptor endocytosis and Smad signaling require synaptojanin1, PI3K-C2α-, and INPP4B-mediated phosphoinositide conversions.
Aki S, Yoshioka K, Takuwa N, Takuwa Y
Molecular biology of the cell 2020 Mar 1;31(5):360-372
Molecular biology of the cell 2020 Mar 1;31(5):360-372
Physiological blood-brain transport is impaired with age by a shift in transcytosis.
Yang AC, Stevens MY, Chen MB, Lee DP, Stähli D, Gate D, Contrepois K, Chen W, Iram T, Zhang L, Vest RT, Chaney A, Lehallier B, Olsson N, du Bois H, Hsieh R, Cropper HC, Berdnik D, Li L, Wang EY, Traber GM, Bertozzi CR, Luo J, Snyder MP, Elias JE, Quake SR, James ML, Wyss-Coray T
Nature 2020 Jul;583(7816):425-430
Nature 2020 Jul;583(7816):425-430
HHV-7 U21 exploits Golgi quality control carriers to reroute class I MHC molecules to lysosomes.
Dirck AT, Whyte ML, Hudson AW
Molecular biology of the cell 2020 Feb 1;31(3):196-208
Molecular biology of the cell 2020 Feb 1;31(3):196-208
Structurally distinct endocytic pathways for B cell receptors in B lymphocytes.
Roberts AD, Davenport TM, Dickey AM, Ahn R, Sochacki KA, Taraska JW
Molecular biology of the cell 2020 Dec 1;31(25):2826-2840
Molecular biology of the cell 2020 Dec 1;31(25):2826-2840
Acute Statin Treatment Improves Antibody Accumulation in EGFR- and PSMA-Expressing Tumors.
Pereira PMR, Mandleywala K, Ragupathi A, Lewis JS
Clinical cancer research : an official journal of the American Association for Cancer Research 2020 Dec 1;26(23):6215-6229
Clinical cancer research : an official journal of the American Association for Cancer Research 2020 Dec 1;26(23):6215-6229
A unique role for clathrin light chain A in cell spreading and migration.
Tsygankova OM, Keen JH
Journal of cell science 2019 May 15;132(10)
Journal of cell science 2019 May 15;132(10)
Chlamydia exploits filopodial capture and a macropinocytosis-like pathway for host cell entry.
Ford C, Nans A, Boucrot E, Hayward RD
PLoS pathogens 2018 May;14(5):e1007051
PLoS pathogens 2018 May;14(5):e1007051
Rare Disease Mechanisms Identified by Genealogical Proteomics of Copper Homeostasis Mutant Pedigrees.
Zlatic SA, Vrailas-Mortimer A, Gokhale A, Carey LJ, Scott E, Burch R, McCall MM, Rudin-Rush S, Davis JB, Hartwig C, Werner E, Li L, Petris M, Faundez V
Cell systems 2018 Mar 28;6(3):368-380.e6
Cell systems 2018 Mar 28;6(3):368-380.e6
A Flat BAR Protein Promotes Actin Polymerization at the Base of Clathrin-Coated Pits.
Almeida-Souza L, Frank RAW, García-Nafría J, Colussi A, Gunawardana N, Johnson CM, Yu M, Howard G, Andrews B, Vallis Y, McMahon HT
Cell 2018 Jul 12;174(2):325-337.e14
Cell 2018 Jul 12;174(2):325-337.e14
ELMOD1 Stimulates ARF6-GTP Hydrolysis to Stabilize Apical Structures in Developing Vestibular Hair Cells.
Krey JF, Dumont RA, Wilmarth PA, David LL, Johnson KR, Barr-Gillespie PG
The Journal of neuroscience : the official journal of the Society for Neuroscience 2018 Jan 24;38(4):843-857
The Journal of neuroscience : the official journal of the Society for Neuroscience 2018 Jan 24;38(4):843-857
An Aberrant Phosphorylation of Amyloid Precursor Protein Tyrosine Regulates Its Trafficking and the Binding to the Clathrin Endocytic Complex in Neural Stem Cells of Alzheimer's Disease Patients.
Poulsen ET, Iannuzzi F, Rasmussen HF, Maier TJ, Enghild JJ, Jørgensen AL, Matrone C
Frontiers in molecular neuroscience 2017;10:59
Frontiers in molecular neuroscience 2017;10:59
Surface Toll-like receptor 3 expression in metastatic intestinal epithelial cells induces inflammatory cytokine production and promotes invasiveness.
Bugge M, Bergstrom B, Eide OK, Solli H, Kjønstad IF, Stenvik J, Espevik T, Nilsen NJ
The Journal of biological chemistry 2017 Sep 15;292(37):15408-15425
The Journal of biological chemistry 2017 Sep 15;292(37):15408-15425
Influenza virus genome reaches the plasma membrane via a modified endoplasmic reticulum and Rab11-dependent vesicles.
de Castro Martin IF, Fournier G, Sachse M, Pizarro-Cerda J, Risco C, Naffakh N
Nature communications 2017 Nov 9;8(1):1396
Nature communications 2017 Nov 9;8(1):1396
Flat clathrin lattices are dynamic actin-controlled hubs for clathrin-mediated endocytosis and signalling of specific receptors.
Leyton-Puig D, Isogai T, Argenzio E, van den Broek B, Klarenbeek J, Janssen H, Jalink K, Innocenti M
Nature communications 2017 Jul 13;8:16068
Nature communications 2017 Jul 13;8:16068
Nanoscale manipulation of membrane curvature for probing endocytosis in live cells.
Zhao W, Hanson L, Lou HY, Akamatsu M, Chowdary PD, Santoro F, Marks JR, Grassart A, Drubin DG, Cui Y, Cui B
Nature nanotechnology 2017 Aug;12(8):750-756
Nature nanotechnology 2017 Aug;12(8):750-756
Endocytic Pathways Used by Andes Virus to Enter Primary Human Lung Endothelial Cells.
Chiang CF, Flint M, Lin JS, Spiropoulou CF
PloS one 2016;11(10):e0164768
PloS one 2016;11(10):e0164768
madSTORM: a superresolution technique for large-scale multiplexing at single-molecule accuracy.
Yi J, Manna A, Barr VA, Hong J, Neuman KC, Samelson LE
Molecular biology of the cell 2016 Nov 7;27(22):3591-3600
Molecular biology of the cell 2016 Nov 7;27(22):3591-3600
Caveolin- and clathrin-independent entry of BKPyV into primary human proximal tubule epithelial cells.
Zhao L, Marciano AT, Rivet CR, Imperiale MJ
Virology 2016 May;492:66-72
Virology 2016 May;492:66-72
Synaptotagmin-11 inhibits clathrin-mediated and bulk endocytosis.
Wang C, Wang Y, Hu M, Chai Z, Wu Q, Huang R, Han W, Zhang CX, Zhou Z
EMBO reports 2016 Jan;17(1):47-63
EMBO reports 2016 Jan;17(1):47-63
High affinity receptor labeling based on basic leucine zipper domain peptides conjugated with pH-sensitive fluorescent dye: Visualization of AMPA-type glutamate receptor endocytosis in living neurons.
Hayashi A, Asanuma D, Kamiya M, Urano Y, Okabe S
Neuropharmacology 2016 Jan;100:66-75
Neuropharmacology 2016 Jan;100:66-75
Diverse functions of myosin VI elucidated by an isoform-specific α-helix domain.
Wollscheid HP, Biancospino M, He F, Magistrati E, Molteni E, Lupia M, Soffientini P, Rottner K, Cavallaro U, Pozzoli U, Mapelli M, Walters KJ, Polo S
Nature structural & molecular biology 2016 Apr;23(4):300-308
Nature structural & molecular biology 2016 Apr;23(4):300-308
Axial superresolution via multiangle TIRF microscopy with sequential imaging and photobleaching.
Fu Y, Winter PW, Rojas R, Wang V, McAuliffe M, Patterson GH
Proceedings of the National Academy of Sciences of the United States of America 2016 Apr 19;113(16):4368-73
Proceedings of the National Academy of Sciences of the United States of America 2016 Apr 19;113(16):4368-73
Central role for PICALM in amyloid-β blood-brain barrier transcytosis and clearance.
Zhao Z, Sagare AP, Ma Q, Halliday MR, Kong P, Kisler K, Winkler EA, Ramanathan A, Kanekiyo T, Bu G, Owens NC, Rege SV, Si G, Ahuja A, Zhu D, Miller CA, Schneider JA, Maeda M, Maeda T, Sugawara T, Ichida JK, Zlokovic BV
Nature neuroscience 2015 Jul;18(7):978-87
Nature neuroscience 2015 Jul;18(7):978-87
Endophilin marks and controls a clathrin-independent endocytic pathway.
Boucrot E, Ferreira AP, Almeida-Souza L, Debard S, Vallis Y, Howard G, Bertot L, Sauvonnet N, McMahon HT
Nature 2015 Jan 22;517(7535):460-5
Nature 2015 Jan 22;517(7535):460-5
Lack of CD2AP disrupts Glut4 trafficking and attenuates glucose uptake in podocytes.
Tolvanen TA, Dash SN, Polianskyte-Prause Z, Dumont V, Lehtonen S
Journal of cell science 2015 Dec 15;128(24):4588-600
Journal of cell science 2015 Dec 15;128(24):4588-600
Uptake of Helicobacter pylori vesicles is facilitated by clathrin-dependent and clathrin-independent endocytic pathways.
Olofsson A, Nygård Skalman L, Obi I, Lundmark R, Arnqvist A
mBio 2014 May 20;5(3):e00979-14
mBio 2014 May 20;5(3):e00979-14
Characterization of human astrovirus cell entry.
Méndez E, Muñoz-Yañez C, Sánchez-San Martín C, Aguirre-Crespo G, Baños-Lara Mdel R, Gutierrez M, Espinosa R, Acevedo Y, Arias CF, López S
Journal of virology 2014 Mar;88(5):2452-60
Journal of virology 2014 Mar;88(5):2452-60
The X-linked mental retardation protein OPHN1 interacts with Homer1b/c to control spine endocytic zone positioning and expression of synaptic potentiation.
Nakano-Kobayashi A, Tai Y, Nadif Kasri N, Van Aelst L
The Journal of neuroscience : the official journal of the Society for Neuroscience 2014 Jun 25;34(26):8665-71
The Journal of neuroscience : the official journal of the Society for Neuroscience 2014 Jun 25;34(26):8665-71
Functional coupling of chloride-proton exchanger ClC-5 to gastric H+,K+-ATPase.
Takahashi Y, Fujii T, Fujita K, Shimizu T, Higuchi T, Tabuchi Y, Sakamoto H, Naito I, Manabe K, Uchida S, Sasaki S, Ikari A, Tsukada K, Sakai H
Biology open 2014 Jan 15;3(1):12-21
Biology open 2014 Jan 15;3(1):12-21
Alternative endocytosis pathway for productive entry of hepatitis C virus.
Matsuda M, Suzuki R, Kataoka C, Watashi K, Aizaki H, Kato N, Matsuura Y, Suzuki T, Wakita T
The Journal of general virology 2014 Dec;95(Pt 12):2658-2667
The Journal of general virology 2014 Dec;95(Pt 12):2658-2667
Stabilization of actin bundles by a dynamin 1/cortactin ring complex is necessary for growth cone filopodia.
Yamada H, Abe T, Satoh A, Okazaki N, Tago S, Kobayashi K, Yoshida Y, Oda Y, Watanabe M, Tomizawa K, Matsui H, Takei K
The Journal of neuroscience : the official journal of the Society for Neuroscience 2013 Mar 6;33(10):4514-26
The Journal of neuroscience : the official journal of the Society for Neuroscience 2013 Mar 6;33(10):4514-26
Netrin-1-induced activation of Notch signaling mediates glioblastoma cell invasion.
Ylivinkka I, Hu Y, Chen P, Rantanen V, Hautaniemi S, Nyman TA, Keski-Oja J, Hyytiäinen M
Journal of cell science 2013 Jun 1;126(Pt 11):2459-69
Journal of cell science 2013 Jun 1;126(Pt 11):2459-69
The LXR-IDOL axis defines a clathrin-, caveolae-, and dynamin-independent endocytic route for LDLR internalization and lysosomal degradation.
Sorrentino V, Nelson JK, Maspero E, Marques ARA, Scheer L, Polo S, Zelcer N
Journal of lipid research 2013 Aug;54(8):2174-2184
Journal of lipid research 2013 Aug;54(8):2174-2184
The PDZ protein GIPC regulates trafficking of the LPA1 receptor from APPL signaling endosomes and attenuates the cell's response to LPA.
Varsano T, Taupin V, Guo L, Baterina OY Jr, Farquhar MG
PloS one 2012;7(11):e49227
PloS one 2012;7(11):e49227
Cell surface ceramide controls translocation of transferrin receptor to clathrin-coated pits.
Abdel Shakor AB, Atia MM, Kwiatkowska K, Sobota A
Cellular signalling 2012 Mar;24(3):677-84
Cellular signalling 2012 Mar;24(3):677-84
Synaptotagmin 1 is necessary for the Ca2+ dependence of clathrin-mediated endocytosis.
Yao LH, Rao Y, Varga K, Wang CY, Xiao P, Lindau M, Gong LW
The Journal of neuroscience : the official journal of the Society for Neuroscience 2012 Mar 14;32(11):3778-85
The Journal of neuroscience : the official journal of the Society for Neuroscience 2012 Mar 14;32(11):3778-85
Adaptor protein 2-mediated endocytosis of the β-secretase BACE1 is dispensable for amyloid precursor protein processing.
Prabhu Y, Burgos PV, Schindler C, Farías GG, Magadán JG, Bonifacino JS
Molecular biology of the cell 2012 Jun;23(12):2339-51
Molecular biology of the cell 2012 Jun;23(12):2339-51
Rift Valley fever virus strain MP-12 enters mammalian host cells via caveola-mediated endocytosis.
Harmon B, Schudel BR, Maar D, Kozina C, Ikegami T, Tseng CT, Negrete OA
Journal of virology 2012 Dec;86(23):12954-70
Journal of virology 2012 Dec;86(23):12954-70
Multi-colour direct STORM with red emitting carbocyanines.
Lampe A, Haucke V, Sigrist SJ, Heilemann M, Schmoranzer J
Biology of the cell 2012 Apr;104(4):229-37
Biology of the cell 2012 Apr;104(4):229-37
Sphingomyelin synthase 1-generated sphingomyelin plays an important role in transferrin trafficking and cell proliferation.
Shakor ABA, Taniguchi M, Kitatani K, Hashimoto M, Asano S, Hayashi A, Nomura K, Bielawski J, Bielawska A, Watanabe K, Kobayashi T, Igarashi Y, Umehara H, Takeya H, Okazaki T
The Journal of biological chemistry 2011 Oct 14;286(41):36053-36062
The Journal of biological chemistry 2011 Oct 14;286(41):36053-36062
Rapid and efficient clathrin-mediated endocytosis revealed in genome-edited mammalian cells.
Doyon JB, Zeitler B, Cheng J, Cheng AT, Cherone JM, Santiago Y, Lee AH, Vo TD, Doyon Y, Miller JC, Paschon DE, Zhang L, Rebar EJ, Gregory PD, Urnov FD, Drubin DG
Nature cell biology 2011 Mar;13(3):331-7
Nature cell biology 2011 Mar;13(3):331-7
Fast, three-dimensional super-resolution imaging of live cells.
Jones SA, Shim SH, He J, Zhuang X
Nature methods 2011 Jun;8(6):499-508
Nature methods 2011 Jun;8(6):499-508
Intracellular trafficking of hyaluronic acid-chitosan oligomer-based nanoparticles in cultured human ocular surface cells.
Contreras-Ruiz L, de la Fuente M, Párraga JE, López-García A, Fernández I, Seijo B, Sánchez A, Calonge M, Diebold Y
Molecular vision 2011 Jan 27;17:279-90
Molecular vision 2011 Jan 27;17:279-90
Role of the clathrin terminal domain in regulating coated pit dynamics revealed by small molecule inhibition.
von Kleist L, Stahlschmidt W, Bulut H, Gromova K, Puchkov D, Robertson MJ, MacGregor KA, Tomilin N, Pechstein A, Chau N, Chircop M, Sakoff J, von Kries JP, Saenger W, Kräusslich HG, Shupliakov O, Robinson PJ, McCluskey A, Haucke V
Cell 2011 Aug 5;146(3):471-84
Cell 2011 Aug 5;146(3):471-84
Coupling between clathrin-dependent endocytic budding and F-BAR-dependent tubulation in a cell-free system.
Wu M, Huang B, Graham M, Raimondi A, Heuser JE, Zhuang X, De Camilli P
Nature cell biology 2010 Sep;12(9):902-8
Nature cell biology 2010 Sep;12(9):902-8
Ebola virus uses clathrin-mediated endocytosis as an entry pathway.
Bhattacharyya S, Warfield KL, Ruthel G, Bavari S, Aman MJ, Hope TJ
Virology 2010 May 25;401(1):18-28
Virology 2010 May 25;401(1):18-28
EphA8-ephrinA5 signaling and clathrin-mediated endocytosis is regulated by Tiam-1, a Rac-specific guanine nucleotide exchange factor.
Yoo S, Shin J, Park S
Molecules and cells 2010 Jun;29(6):603-9
Molecules and cells 2010 Jun;29(6):603-9
Ornithine decarboxylase antizyme inhibitor 2 regulates intracellular vesicle trafficking.
Kanerva K, Mäkitie LT, Bäck N, Andersson LC
Experimental cell research 2010 Jul 1;316(11):1896-906
Experimental cell research 2010 Jul 1;316(11):1896-906
Disruption of zebrafish cyclin G-associated kinase (GAK) function impairs the expression of Notch-dependent genes during neurogenesis and causes defects in neuronal development.
Bai T, Seebald JL, Kim KE, Ding HM, Szeto DP, Chang HC
BMC developmental biology 2010 Jan 18;10:7
BMC developmental biology 2010 Jan 18;10:7
Assembly of connexin43 into gap junctions is regulated differentially by E-cadherin and N-cadherin in rat liver epithelial cells.
Govindarajan R, Chakraborty S, Johnson KE, Falk MM, Wheelock MJ, Johnson KR, Mehta PP
Molecular biology of the cell 2010 Dec;21(23):4089-107
Molecular biology of the cell 2010 Dec;21(23):4089-107
Rapid turnover of spinules at synaptic terminals.
Tao-Cheng JH, Dosemeci A, Gallant PE, Miller S, Galbraith JA, Winters CA, Azzam R, Reese TS
Neuroscience 2009 Apr 21;160(1):42-50
Neuroscience 2009 Apr 21;160(1):42-50
Phosphoinositide 3-kinase p110beta activity: key role in metabolism and mammary gland cancer but not development.
Ciraolo E, Iezzi M, Marone R, Marengo S, Curcio C, Costa C, Azzolino O, Gonella C, Rubinetto C, Wu H, Dastrù W, Martin EL, Silengo L, Altruda F, Turco E, Lanzetti L, Musiani P, Rückle T, Rommel C, Backer JM, Forni G, Wymann MP, Hirsch E
Science signaling 2008 Sep 9;1(36):ra3
Science signaling 2008 Sep 9;1(36):ra3
Hrs and SNX3 functions in sorting and membrane invagination within multivesicular bodies.
Pons V, Luyet PP, Morel E, Abrami L, van der Goot FG, Parton RG, Gruenberg J
PLoS biology 2008 Sep 2;6(9):e214
PLoS biology 2008 Sep 2;6(9):e214
Mass spectrometry analysis of the native protein complex containing actinin-4 in prostate cancer cells.
Hara T, Honda K, Shitashige M, Ono M, Matsuyama H, Naito K, Hirohashi S, Yamada T
Molecular & cellular proteomics : MCP 2007 Mar;6(3):479-91
Molecular & cellular proteomics : MCP 2007 Mar;6(3):479-91
A role for clathrin in reassembly of the Golgi apparatus.
Radulescu AE, Siddhanta A, Shields D
Molecular biology of the cell 2007 Jan;18(1):94-105
Molecular biology of the cell 2007 Jan;18(1):94-105
Role of internalization of M2 muscarinic receptor via clathrin-coated vesicles in desensitization of the muscarinic K+ current in heart.
Yamanushi TT, Shui Z, Leach RN, Dobrzynski H, Claydon TW, Boyett MR
American journal of physiology. Heart and circulatory physiology 2007 Apr;292(4):H1737-46
American journal of physiology. Heart and circulatory physiology 2007 Apr;292(4):H1737-46
GBF1, a cis-Golgi and VTCs-localized ARF-GEF, is implicated in ER-to-Golgi protein traffic.
Zhao X, Claude A, Chun J, Shields DJ, Presley JF, Melançon P
Journal of cell science 2006 Sep 15;119(Pt 18):3743-53
Journal of cell science 2006 Sep 15;119(Pt 18):3743-53
Interaction of SPIN90 with syndapin is implicated in clathrin-mediated endocytic pathway in fibroblasts.
Kim SH, Choi HJ, Lee KW, Hong NH, Sung BH, Choi KY, Kim SM, Chang S, Eom SH, Song WK
Genes to cells : devoted to molecular & cellular mechanisms 2006 Oct;11(10):1197-211
Genes to cells : devoted to molecular & cellular mechanisms 2006 Oct;11(10):1197-211
Trimerisation is important for the function of clathrin at the mitotic spindle.
Royle SJ, Lagnado L
Journal of cell science 2006 Oct 1;119(Pt 19):4071-8
Journal of cell science 2006 Oct 1;119(Pt 19):4071-8
Epidermal growth factor receptor exposed to oxidative stress undergoes Src- and caveolin-1-dependent perinuclear trafficking.
Khan EM, Heidinger JM, Levy M, Lisanti MP, Ravid T, Goldkorn T
The Journal of biological chemistry 2006 May 19;281(20):14486-93
The Journal of biological chemistry 2006 May 19;281(20):14486-93
ST6Gal-I restrains CD22-dependent antigen receptor endocytosis and Shp-1 recruitment in normal and pathogenic immune signaling.
Grewal PK, Boton M, Ramirez K, Collins BE, Saito A, Green RS, Ohtsubo K, Chui D, Marth JD
Molecular and cellular biology 2006 Jul;26(13):4970-81
Molecular and cellular biology 2006 Jul;26(13):4970-81
Vascular endothelial cadherin controls VEGFR-2 internalization and signaling from intracellular compartments.
Lampugnani MG, Orsenigo F, Gagliani MC, Tacchetti C, Dejana E
The Journal of cell biology 2006 Aug 14;174(4):593-604
The Journal of cell biology 2006 Aug 14;174(4):593-604
SPG3A protein atlastin-1 is enriched in growth cones and promotes axon elongation during neuronal development.
Zhu PP, Soderblom C, Tao-Cheng JH, Stadler J, Blackstone C
Human molecular genetics 2006 Apr 15;15(8):1343-53
Human molecular genetics 2006 Apr 15;15(8):1343-53
Exchange of clathrin, AP2 and epsin on clathrin-coated pits in permeabilized tissue culture cells.
Yim YI, Scarselletta S, Zang F, Wu X, Lee DW, Kang YS, Eisenberg E, Greene LE
Journal of cell science 2005 Jun 1;118(Pt 11):2405-13
Journal of cell science 2005 Jun 1;118(Pt 11):2405-13
Analysis of foot-and-mouth disease virus internalization events in cultured cells.
O'Donnell V, LaRocco M, Duque H, Baxt B
Journal of virology 2005 Jul;79(13):8506-18
Journal of virology 2005 Jul;79(13):8506-18
Clathrin-independent endocytosis of ubiquitinated cargos.
Sigismund S, Woelk T, Puri C, Maspero E, Tacchetti C, Transidico P, Di Fiore PP, Polo S
Proceedings of the National Academy of Sciences of the United States of America 2005 Feb 22;102(8):2760-5
Proceedings of the National Academy of Sciences of the United States of America 2005 Feb 22;102(8):2760-5
TTP specifically regulates the internalization of the transferrin receptor.
Tosoni D, Puri C, Confalonieri S, Salcini AE, De Camilli P, Tacchetti C, Di Fiore PP
Cell 2005 Dec 2;123(5):875-88
Cell 2005 Dec 2;123(5):875-88
A novel GTPase-activating protein for ARF6 directly interacts with clathrin and regulates clathrin-dependent endocytosis.
Tanabe K, Torii T, Natsume W, Braesch-Andersen S, Watanabe T, Satake M
Molecular biology of the cell 2005 Apr;16(4):1617-28
Molecular biology of the cell 2005 Apr;16(4):1617-28
Increased expression of protein C-mannosylation in the aortic vessels of diabetic Zucker rats.
Ihara Y, Manabe S, Kanda M, Kawano H, Nakayama T, Sekine I, Kondo T, Ito Y
Glycobiology 2005 Apr;15(4):383-92
Glycobiology 2005 Apr;15(4):383-92
Actin dynamics coupled to clathrin-coated vesicle formation at the trans-Golgi network.
Carreno S, Engqvist-Goldstein AE, Zhang CX, McDonald KL, Drubin DG
The Journal of cell biology 2004 Jun 21;165(6):781-8
The Journal of cell biology 2004 Jun 21;165(6):781-8
Endocytosis of epithelial apical junctional proteins by a clathrin-mediated pathway into a unique storage compartment.
Ivanov AI, Nusrat A, Parkos CA
Molecular biology of the cell 2004 Jan;15(1):176-88
Molecular biology of the cell 2004 Jan;15(1):176-88
Differences in endosomal targeting of human (beta)1- and (beta)2-adrenergic receptors following clathrin-mediated endocytosis.
Liang W, Curran PK, Hoang Q, Moreland RT, Fishman PH
Journal of cell science 2004 Feb 15;117(Pt 5):723-34
Journal of cell science 2004 Feb 15;117(Pt 5):723-34
Detergent-resistant membrane microdomains facilitate Ib oligomer formation and biological activity of Clostridium perfringens iota-toxin.
Hale ML, Marvaud JC, Popoff MR, Stiles BG
Infection and immunity 2004 Apr;72(4):2186-93
Infection and immunity 2004 Apr;72(4):2186-93
RNAi-mediated Hip1R silencing results in stable association between the endocytic machinery and the actin assembly machinery.
Engqvist-Goldstein AE, Zhang CX, Carreno S, Barroso C, Heuser JE, Drubin DG
Molecular biology of the cell 2004 Apr;15(4):1666-79
Molecular biology of the cell 2004 Apr;15(4):1666-79
Targeted chemical disruption of clathrin function in living cells.
Moskowitz HS, Heuser J, McGraw TE, Ryan TA
Molecular biology of the cell 2003 Nov;14(11):4437-47
Molecular biology of the cell 2003 Nov;14(11):4437-47
PP1 inhibitor induces degradation of RETMEN2A and RETMEN2B oncoproteins through proteosomal targeting.
Carniti C, Perego C, Mondellini P, Pierotti MA, Bongarzone I
Cancer research 2003 May 1;63(9):2234-43
Cancer research 2003 May 1;63(9):2234-43
Spatial regulation of Galphai protein signaling in clathrin-coated membrane microdomains containing GAIP.
Elenko E, Fischer T, Niesman I, Harding T, McQuistan T, Von Zastrow M, Farquhar MG
Molecular pharmacology 2003 Jul;64(1):11-20
Molecular pharmacology 2003 Jul;64(1):11-20
Sustained stimulation shifts the mechanism of endocytosis from dynamin-1-dependent rapid endocytosis to clathrin- and dynamin-2-mediated slow endocytosis in chromaffin cells.
Artalejo CR, Elhamdani A, Palfrey HC
Proceedings of the National Academy of Sciences of the United States of America 2002 Apr 30;99(9):6358-63
Proceedings of the National Academy of Sciences of the United States of America 2002 Apr 30;99(9):6358-63
HIP1 functions in clathrin-mediated endocytosis through binding to clathrin and adaptor protein 2.
Metzler M, Legendre-Guillemin V, Gan L, Chopra V, Kwok A, McPherson PS, Hayden MR
The Journal of biological chemistry 2001 Oct 19;276(42):39271-6
The Journal of biological chemistry 2001 Oct 19;276(42):39271-6
Dynamin is involved in human epithelial cell vacuolation caused by the Helicobacter pylori-produced cytotoxin VacA.
Suzuki J, Ohnsihi H, Shibata H, Wada A, Hirayama T, Iiri T, Ueda N, Kanamaru C, Tsuchida T, Mashima H, Yasuda H, Fujita T
The Journal of clinical investigation 2001 Feb;107(3):363-70
The Journal of clinical investigation 2001 Feb;107(3):363-70
Hsc70 chaperones clathrin and primes it to interact with vesicle membranes.
Jiang R, Gao B, Prasad K, Greene LE, Eisenberg E
The Journal of biological chemistry 2000 Mar 24;275(12):8439-47
The Journal of biological chemistry 2000 Mar 24;275(12):8439-47
Role of cyclin G-associated kinase in uncoating clathrin-coated vesicles from non-neuronal cells.
Greener T, Zhao X, Nojima H, Eisenberg E, Greene LE
The Journal of biological chemistry 2000 Jan 14;275(2):1365-70
The Journal of biological chemistry 2000 Jan 14;275(2):1365-70
Clathrin-coated vesicles bearing GAIP possess GTPase-activating protein activity in vitro.
Fischer T, Elenko E, McCaffery JM, DeVries L, Farquhar MG
Proceedings of the National Academy of Sciences of the United States of America 1999 Jun 8;96(12):6722-7
Proceedings of the National Academy of Sciences of the United States of America 1999 Jun 8;96(12):6722-7
Overexpression of proteins containing tyrosine- or leucine-based sorting signals affects transferrin receptor trafficking.
Nordeng TW, Bakke O
The Journal of biological chemistry 1999 Jul 23;274(30):21139-48
The Journal of biological chemistry 1999 Jul 23;274(30):21139-48
An actin-binding protein of the Sla2/Huntingtin interacting protein 1 family is a novel component of clathrin-coated pits and vesicles.
Engqvist-Goldstein AE, Kessels MM, Chopra VS, Hayden MR, Drubin DG
The Journal of cell biology 1999 Dec 27;147(7):1503-18
The Journal of cell biology 1999 Dec 27;147(7):1503-18
Characterization of the adaptor-related protein complex, AP-3.
Simpson F, Peden AA, Christopoulou L, Robinson MS
The Journal of cell biology 1997 May 19;137(4):835-45
The Journal of cell biology 1997 May 19;137(4):835-45
Targeting and mistargeting of plasma membrane adaptors in vitro.
Seaman MN, Ball CL, Robinson MS
The Journal of cell biology 1993 Dec;123(5):1093-105
The Journal of cell biology 1993 Dec;123(5):1093-105
100-kD proteins of Golgi- and trans-Golgi network-associated coated vesicles have related but distinct membrane binding properties.
Wong DH, Brodsky FM
The Journal of cell biology 1992 Jun;117(6):1171-9
The Journal of cell biology 1992 Jun;117(6):1171-9
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- Western blot analysis of Clathrin, Heavy chain was performed by loading 25 µg of human brain lysates onto an SDS polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked at 4ºC overnight. The membrane was probed with a Clathrin, Heavy chain monoclonal antibody (Product # MA1-065) at a dilution of 1:300 overnight at 4°C, washed in TBST, and probed with an HRP-conjugated goat anti-mouse IgG + IgM (H+L) cross-adsorbed secondary antibody for 1 hr at room temperature in the dark. Chemiluminescent detection was performed using Pierce ECL Plus Western Blotting Substrate (Product # 32132). Results show a band at ~180 kDa.
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- Knockdown of Clathrin Heavy Chain was achieved by transfecting HeLa with Clathrin Heavy Chain specific siRNAs (Silencer® select Product # S475, S477). Western blot analysis (Fig. a) was performed using Whole cell extracts from the Clathrin Heavy Chain knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with Clathrin Heavy Chain Monoclonal Antibody (X22) (Product # MA1-065, 1:500 ) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to Clathrin Heavy Chain.
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- Western blot was performed using Anti-Clathrin Heavy Chain Monoclonal Antibody (X22) (Product # MA1-065) and a 191kDa band corresponding to Clathrin Heavy Chain was observed across cell lines. Whole cell extracts (30 µg lysate) of MCF7 (Lane 1), U-87 MG (Lane 2), HeLa (Lane 3) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23002) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:500) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
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- Immunofluorescent analysis of Clathrin, Heavy chain using Anti-Clathrin, Heavy chain Monoclonal Antibody (X22) (Product # MA1-065) shows staining in Hela Cells. Clathrin, Heavy chain staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Clathrin, Heavy chain (Product # MA1-065) at a dilution of 1:200 over night at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody (Product # 35503, Goat Anti-Mouse). Images were taken at 60X magnification.
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- Immunofluorescent analysis of Clathrin, Heavy chain using Anti-Clathrin, Heavy chain Monoclonal Antibody (X22) (Product # MA1-065) shows staining in NCI-H460 Cells. Clathrin, Heavy chain staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Clathrin, Heavy chain (Product # MA1-065) at a dilution of 1:200 over night at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody (Product # 35503, Goat Anti-Mouse). Images were taken at 60X magnification.
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- Immunofluorescent analysis of Clathrin, Heavy chain using Anti-Clathrin, Heavy chain Monoclonal Antibody (X22) (Product # MA1-065) shows staining in U251 Cells. Clathrin, Heavy chain staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with or an antibody recognizing Clathrin, Heavy chain (Product # MA1-065) at a dilution of 1:200 over night at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody (Product # 35503, Goat Anti-Mouse). Images were taken at 60X magnification.
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- Immunohistochemistry was performed on cancer biopsies of deparaffinized Human breast carcinoma tissue. To expose target proteins, heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer, microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:100 with a mouse monoclonal antibody recognizing Clathrin, Heavy chain (Product # MA1-065) or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP, followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.
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- Immunohistochemistry was performed on normal biopsies of deparaffinized Human colon tissue. To expose target proteins, heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer, microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:100 with a mouse monoclonal antibody recognizing Clathrin, Heavy chain (Product # MA1-065) or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP, followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.
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- Figure 2 The extent of Clathrin/APP and AP2/APP colocalization is reduced in NSCs from patients with AD . Confocal microscopy analysis of double staining using rabbit anti-APP and mouse anti-AP2 (A) and mouse anti-Clathrin (B) in control (C18) and AD neurons with PS1 mutations (L286V; M146L; A246E). Colocalization analysis is reported in (C) . The (R) coefficient (Pearson's coefficient) was used for the quantitative and comparative analyses. The extent of colocalization was calculated in at least five separate fields per slide in 10 different slides for each NSCs. The data are expressed as mean +- SEM. Scale bars are 5 mum for APP/AP2 and 6 mum for APP/Clathrin colocalization. Scale bars in high-resolution pictures are 5 mum. Statistically significant differences were calculated by one-way ANOVA and Tukey's post hoc test.
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- Figure 3 Tyr kinase inhibitors restore APP colocalization with Clathrin and AP2 in NSCs from patients with AD . (A) IP analysis of control and PS1 neurons that were exposed, or not exposed, to Sunitinib and PP2. Control C18 and AD samples (LV, ML, AE) were immunoprecipitated with anti-pTyr agarose conjugated antibody (4G10) and analyzed with rabbit anti-APP (clone Y188). The image is representative of four independent experiments. Quantification is reported in (B) . Data were normalized with pTyr pulled down levels (input) and expressed as % of C18. Statistically significant differences were calculated by one-way ANOVA for repeated measures followed by Tukey's post hoc test for multiple comparisons. Confocal microscopy analysis of double staining using rabbit anti-APP and mouse anti-AP2 (C) and mouse anti-Clathrin (D) in controls and in neurons carrying L286V or M146L mutation on PS1 gene following exposure to the Tyr kinase inhibitor Sunitinib. The panels are representative of four different experiments performed in triplicate. (E) reports quantitative analysis of APP colocalization to AP2 and Clathrin after 12 h of exposure to Sunitinib. (E) also reports colocalization analysis after PP2 exposure. The (R) coefficient (Pearson's coefficient) was used for the quantitative and comparative analyses. The extent of colocalization was calculated in five separate fields per slide in five different slides for each experimental point. The data are expressed as mean +- SEM.
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- Figure 4 Tyr phosphatase inhibitors reduce APP colocalization with Clathrin and AP2 and increase phosphorylation of APP Tyr in control neurons . Confocal microscopy analysis of double staining with rabbit anti-APP and mouse anti-AP2 (A) or mouse anti-Clathrin (B) in C18, L286V, and M146L neurons before and after Tyr phosphatase inhibitor (TC2153) exposure. The panels are representative of five different experiments performed in duplicate. (C) Colocalization analysis of APP to AP2 and Clathrin following incubation with TC2153 and BVT948 (BVT) inhibitors in C18 neurons and in AD neurons. The (R) coefficient (Pearson's coefficient) was used for the quantitative and comparative analyses. The extent of colocalization was calculated in five separate fields per slide in four different slides for each experimental point. The data are expressed as mean +- SEM. Scale bars 6 mum and 4 mum. Statistically significant differences were calculated by one-way ANOVA and Tukey's post hoc test. (D,E) . IP analysis of C18 (D) and AD neurons (E) before and after exposure to TC2153 and BVT948 (TC, BVT). Samples were immunoprecipitated with anti-pTyr agarose conjugated antibody (4G10) and analyzed with rabbit anti-APP (clone Y188). Densitometric analysis is reported in (F) . Data were normalized with IgG levels and expressed as % of C18. Statistically significant differences were calculated using Student's t -test.
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- Figure 5 APP/AP2 and APP/Clathrin binding analysis in cortical tissues and fibroblasts of Gottingen minipigs with a PS1 M146I mutation in the PS1 gene . (A) CoIP analysis of cortical tissues and fibroblasts from control (Ctrl) and PS1 M146I minipigs. Samples were immunoprecipitated with rabbit anti-APP and analyzed with mouse anti-Clathrin (Clath) and mouse anti-AP2. (B) WB analysis of total lysate from controls (WT and Ctrl) and PS1 M146I minipigs. Densitometric analysis is reported in (C) . Data were normalized to the corresponding beta-actin values and expressed as % of Ctrl. Data from CoIP samples were normalized to the corresponding APP input amount and expressed as % of Ctrl. Statistically significant differences were calculated using Student's t -test. (D,E) Confocal microscopy analysis of double staining with rabbit anti-APP and mouse anti-AP2 (D) and mouse anti-Clathrin (E) in fibroblasts from Ctrl and PS1M146I minipigs in the presence of TC2153 or Sunitinib. The panels are representative of five different experiments performed in duplicate in Ctrl and PS1 fibroblasts from two independent control (Ctrl 1-2) and three PS1 (PS1a-c) minipigs. Colocalization analysis is reported in (F) . The (R) coefficient (Pearson's coefficient) was used for the quantitative and comparative analyses. The extent of colocalization was calculated in six separate fields per slide in four different slides for each cell line. Scale bars 1 muM and 5 muM in high magnification. Statistically si
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- Figure 1 FCHSD2 Is a Bona Fide CME Protein Responsible for a Major Fraction of the ARP2/3 Contribution to CME (A) Top: Scheme showing the domain organization of FCHSD proteins. Bottom: Immunoblots for N-WASP and Intersectin1 (ITSN1) from pull down experiments from brain extracts using GST-tagged FCHSD1 and FCHSD2 SH3 domains. Lower portion shows Coomassie staining of baits. (B) Immunofluorescence showing colocalization between endogenous FCHSD2 and clathrin heavy chain. (C) TIRF image showing colocalization of FCHSD2 and clathrin. HeLa cells stably expressing FCHSD2-Venus and transfected with mCherry-clathrin light chain. (D) Left: Examples of the dynamics of FCHSD2 with different CME proteins. HeLa cells stably expressing FCHSD2-Venus were transfected with mCherry-clathrinLC, FusionRed-ITSN1L, FusionRed-Dynamin1, or mCherry-ARP3 and imaged live by TIRF microscopy. Time zero was set as the peak of FCHSD2 recruitment. Events are pseudocolored to match graphs on the right. Right: Summary graphs for the timing of recruitment of FCHSD2 versus CME proteins (n = 90, 48, 120, and 144 events for FCHSD2/clathrin, FCHSD2/ITSN1L, FCHSD2/Dynamin, and FCHSD2/ARP3, respectively). Full data including error bars are shown in Figure S1 A. (E) Transferrin uptake assay by flow cytometry. Uptake measurements were normalized as described in STAR Methods . Each value represents median fluorescence from at least 5,000 cells (n = 10, mean +- SD). (F) Left: Kymographs of BSC1 AP2sigma2-GFP cells sile
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- Figure 5 FCHSD2 Localizes to the Plasma Membrane Side of CCPs (A) Sequential widefield (WF) and TIRF imaging of endocytic proteins allows the distinction between proteins that stay on the membrane and proteins that move away from the membrane. (B) Results for AP2, dynamin, and FCHSD2 using the experimental paradigm explained in (A). Image series of representative events generated from videos at 0.5 Hz. (n = 54, 51, and 53 events for AP2, dynamin, and FCHSD2 respectively, mean +- SEM). (C-E) Comparative localization of AP2 and FCHSD2 with CCPs by confocal (C), STED (D), and 3D STED (E) microscopy. Stable HeLa cells for AP2sigma2-GFP and FCHSD2-Venus cells were stained with anti-GFP and anti-Clathrin antibodies. Cartoon representations of the views are shown on the right hand side of super resolution images. Scale bars, 0.25 mum. (F) Curvature preference of FCHSD2 BAR by nanoparticle tracking analysis (NTA). Graph showing the size distribution of the total liposome population and the FCHSD2 BAR-sfGFP bound subpopulation. Total population distribution is measured by tracking particles diffracting light while FCHSD2 bound population is measured by tracking particles emitting light from GFP excitation. FCHSD2 BAR-sfGFP added at 1 nM (n = 3 experiments, mean +- SD). (G and H) Single particle cryoEM of F2B1 (FCHSD2 F-BAR+SH3-1). (G) 3D reconstruction from 30,207 particles. (H) The densities at the tip of the 3D map are compatible with an SH3 domain. In magenta is a surface represent
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- Fig 2 Knockdown of selected genes required for ANDV infection. Western blots of indicated proteins after knockdown with gene-specific siRNA against (A) dynamin 2 (DNM2); (B) clathrin heavy chain (CLTC); (C) AP2M1; (D) caveolin 1 (CAV1); (E) CDC42; (F) ARF6; (G) NSF; (H) RAB5C; or (I) TSG101; or non-targeting siRNA transfection control (NT). siRNAs were transfected into HMVEC-L at the concentration of 100 nM for 48 h (see Table 1 for more information regarding these genes). The cells were then infected with ANDV (MOI = 0.5) for 24 h or 48 h. Western blots were performed post infection to ensure knockdown of the specific protein expression. Molecular weight of each specific protein is indicated on the right side of each panel. The blots were also probed with beta-actin specific antibody as the gel-loading control. (A) and (B) show only the results collected at 48 h.
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- Figure 5. CXCL10 production in metastatic IECs is elicited independent of poly(I:C) internalization but requires endosomal acidification. A , confocal microscopy image of HT29 cells treated with poly(I:C) Rhodamine ( red , 5 mug/ml) for 2 h before cells were washed and fixed, and the plasma membrane ( PM ) was stained with an antibody against Na,K-ATPase ( PM A488 , green ). Images ( top ) show co-localization ( Coloc , left ) and single tracks of poly(I:C) Rhodamine ( center ) and plasma membrane PM A488 staining ( right ) of the area denoted by the square in the main image. Scale bar = 5 mum. B and C , SW620 ( B ) or HT29 ( C ) cells were treated with poly(I:C) ( Added Poly(I:C) ) or double-stranded DNA dA:dT ( Added dA:dT ) added in solution or by plating cells in wells precoated with poly(I:C) ( Coated Poly(I:C) ) or dA:dT ( Coated dA:dT ) with the given concentrations for 24 h before CXCL10 release was determined by ELISA. D , CXCL10 production in HT29 cells exposed to HMW poly(I:C) or LMW poly(I:C) (2 mug/ml) either added in solution ( Added ) or by plating cells in wells precoated with poly(I:C) ( Coated ) for 21 h. E , CXCL10 release in HT29 cells pretreated with anti-TLR3 (15 or 5 mug/ml) or control goat IgG (15 mug/ml) for 1 h prior to stimulation with poly(I:C) (5 or 2 mug/ml) for 10 h. CXCL10 content in the supernatant was assessed by ELISA. **, p < 0.01; *, p < 0.05 versus cells pretreated with control IgG (two-way ANOVA, Bonferroni post-test). The results in A-D
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- Experimental details
- Figure 3. Characterization of expansion isotropy using Ten-fold Robust Expansion Microscopy (TREx). ( A ) U2OS knock-in cells with homozygous NUP96-GFP, amplified with anti-GFP antibodies. ( B ) One nucleus from boxed region of ( A ), imaged by confocal microscopy after TREx. ( C ) High-resolution view of several nuclear pores from boxed region (1) of panel ( B ), showing both anti-GFP (magenta) and anti-NUP153 (endogenous nuclear pore protein, cyan) staining. ( D ) High-resolution view of several nuclear pores from boxed region (2) of panel ( B ) (top). Distribution of diameters of individual nuclear pores (bottom), corrected for the macroscopic expansion factor of 9.5x. N = 60 nuclear pore complexes (NPCs) from three spatially separated cells. ( E ) U2OS cells stained for clathrin heavy chain and tubulin, representative line scan over clathrin-coated pit (CCP) showing central . ( F ) Quantification of CCP diameter. Plotted mean +- SD (1.16 +- 0.2 um) of 25 CCPs from five cells (two independent experiments). ( G ) High-resolution view of microtubules in extracted COS7 cell and corresponding line scans with mean peak-to-peak distance indicated. ( H ) Maximum projection of pre-expansion 3D gSTED acquisition (left) and maximum projection of tilescan acquisition (42 tiles, post-expansion size ~750 x 650 um) of the same cell post-expansion (right). ( I ) Post-expansion single field of view, as indicated with magenta box in ( D ), aligned with the pre-expansion image (gray) by