Calcyon, a novel partner of clathrin light chain, stimulates clathrin-mediated endocytosis.
In the central nervous system, clathrin-mediated endocytosis is crucial for efficient synaptic transmission. Clathrin-coated vesicle assembly and disassembly is regulated by some 30 adaptor and accessory proteins, most of which interact with clathrin heavy chain. Using the calcyon cytosolic domain as bait, we isolated clathrin light chain in a yeast two-hybrid screen. The interaction domain was mapped to the heavy chain binding domain and C-terminal regions of light chain. Further, the addition of the calcyon C terminus stimulated clathrin self-assembly in a dose-dependent fashion. Calcyon, which is a single transmembrane protein predominantly expressed in brain, localized to vesicular compartments within pre- and postsynaptic structures. There was a high degree of overlap in the distribution of LC and calcyon in neuronal dendrites, spines, and cell bodies. Co-immunoprecipitation studies further suggested an association of calcyon with the clathrin-mediated endocytic machinery. Compared with controls, HEK293 cells overexpressing calcyon exhibited significantly enhanced transferrin uptake but equivalent levels of recycling. Conversely, transferrin uptake was largely abolished in neocortical neurons obtained from mice homozygous for a calcyon null allele, whereas recycling proceeded at wild type levels. Collectively, these data indicate a role for calcyon in clathrin-mediated endocytosis in brain.
Xiao
Jiping
J
Department of Pharmacology and Toxicology, Medical College of Georgia, Augusta, Georgia 30912, USA.
Dai
Rujuan
R
Negyessy
Laszlo
L
Bergson
Clare
C
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002478:Cells, Cultured; D002490:Central Nervous System / Q000502:physiology; D033941:Clathrin Light Chains / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004705:Endocytosis / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D008253:Macaca mulatta; D008565:Membrane Proteins / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D008854:Microscopy, Electron; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009474:Neurons / Q000378:metabolism; D017397:Prefrontal Cortex / Q000378:metabolism / Q000648:ultrastructure; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009435:Synaptic Transmission / Q000502:physiology; D020798:Two-Hybrid System Techniques
D033941:Clathrin Light Chains; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011994:Recombinant Proteins; C405464:calcyon