title: Recycling of the membrane-anchored chemokine, CX3CL1.
abstract: CX(3)CL1 (fractalkine) plays an important role in inflammation by acting as both chemoattractant and as an adhesion molecule. As for other chemokines, expression of CX(3)CL1 is known to be regulated at the level of transcription and translation. The unique transmembrane structure of CX(3)CL1 raises the possibility of additional functional regulation by altering its abundance at the cell surface. This could be accomplished in principle by changes in traffic between subcellular compartments. To analyze this possibility we examined the subcellular distribution of CX(3)CL1 in human ECV-304 cells stably expressing untagged or green fluorescent protein-tagged forms of the chemokine. CX(3)CL1 was present in two distinct compartments, diffusely on the plasma membrane and in a punctate juxtanuclear compartment. The latter shared some features with, yet was distinct from the conventional endocytic pathway and may represent a specialized recycling subcompartment. Accordingly, surface CX(3)CL1 was found to be in dynamic equilibrium with the juxtanuclear vesicular compartment. Intracellular CX(3)CL1 co-localized with the SNARE (soluble N-ethylmaleimide factor attachment protein receptor) proteins syntaxin-13 and VAMP-3. Cleavage of VAMP-3 by tetanus toxin or impairment of syntaxin-13 function by expression of a dominant-negative allele inhibited the ability of internalized CX(3)CL1 to traffic back to the plasma membrane. These data demonstrate the existence of a dynamic, SNARE-mediated recycling of CX(3)CL1 from the cell surface to and from an endomembrane storage compartment. The intracellular storage depot may serve as a source of the chemokine that could be rapidly mobilized by stimuli.
authors.affiliation: The Hospital for Sick Children Research Institute and the University of Toronto, Canada.
authors.forename: Guang-Ying
authors.initials: GY
authors.lastname: Liu
authors.forename: Vathany
authors.initials: V
authors.lastname: Kulasingam
authors.forename: R Todd
authors.initials: RT
authors.lastname: Alexander
authors.forename: Nicolas
authors.initials: N
authors.lastname: Touret
authors.forename: Alan M
authors.initials: AM
authors.lastname: Fong
authors.forename: Dhavalkumar D
authors.initials: DD
authors.lastname: Patel
authors.forename: Lisa A
authors.initials: LA
authors.lastname: Robinson
mesh_terms: D000818:Animals; D001483:Base Sequence; D001693:Biological Transport, Active; D019556:COS Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D054428:Chemokine CX3CL1; D020523:Chemokines, CX3C / Q000235:genetics / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004247:DNA / Q000235:genetics; D011992:Endosomes / Q000378:metabolism; D036681:Fluorescence Recovery After Photobleaching; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D050600:SNARE Proteins; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection; D033921:Vesicular Transport Proteins / Q000378:metabolism
chemical_list: C515538:CX3CL1 protein, human; D054428:Chemokine CX3CL1; D020523:Chemokines, CX3C; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins; D004247:DNA