Peptide motifs: building the clathrin machinery.
Clathrin-coated vesicles (CCVs) form at the plasma membrane, where they select cargo for endocytic entry into cells, and at the trans-Golgi network (TGN) and the endosomal system, where they generate carrier vesicles that transport proteins between these compartments. We have used subcellular fractionation and tandem mass spectrometry to identify proteins associated with brain CCVs. The resulting proteome contained a near complete inventory of the major functional proteins of synaptic vesicles (SVs), suggesting that clathrin-mediated endocytosis provides a major mechanism to recycle SV membrane proteins following neurotransmitter release. Additionally, we identified several new components of the machineries for clathrin-mediated membrane budding, including enthoprotin/epsinR and NECAP 1/2. These proteins bind with high specificity to the ear domains of the clathrin adaptor proteins (APs)-1 and -2, and, intriguingly, they each utilize novel peptide motifs based around the core sequence ØXXØ. Detailed mutational analysis of these motifs, coupled with structural studies of the ear domains, has revealed the basis of their specificity for clathrin adaptors. Moreover, the motifs have now been recognized in multiple proteins functioning in clathrin-mediated membrane trafficking, revealing new mechanisms in the formation and function of CCVs. Thus, proteomics analysis of isolated organelles can provide insights ranging from peptide motifs to global organelle function.
McPherson
Peter S
PS
Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, QC H3A 2B4, Canada. peter.mcpherson@mcgill.ca
Ritter
Brigitte
B
D000818:Animals; D001665:Binding Sites; D002462:Cell Membrane / Q000502:physiology; D002966:Clathrin / Q000737:chemistry / Q000502:physiology*; D022163:Clathrin-Coated Vesicles / Q000502:physiology*; D011992:Endosomes / Q000502:physiology; D008954:Models, Biological; D008958:Models, Molecular; D009473:Neuronal Plasticity
D002966:Clathrin