Receptor clustering drives polarized assembly of ankyrin.
Expression of the L1 family cell adhesion molecule neuroglian in Drosophila S2 cells leads to cell aggregation and polarized ankyrin accumulation at sites of cell-cell contact. Thus neuroglian adhesion generates a spatial cue for polarized assembly of ankyrin and the spectrin cytoskeleton. Here we characterized a chimera of the extracellular and transmembrane domains of rat CD2 fused to the cytoplasmic domain of neuroglian. The chimera was used to test the hypothesis that clustering of neuroglian at sites of adhesion generates the signal that activates ankyrin binding. Abundant expression of the chimera at the plasma membrane was not a sufficient cue to drive ankyrin assembly, since ankyrin remained diffusely distributed throughout the cytoplasm of CD2-neuroglian-expressing cells. However, ankyrin became highly enriched at sites of antibody-induced capping of CD2-neuroglian. Spectrin codistributed with ankyrin at capped sites. A green fluorescent protein-tagged ankyrin was used to monitor ankyrin distribution in living cells. Enhanced green fluorescent protein-ankyrin behaved identically to antibody-stained endogenous ankyrin, proving that the polarized accumulation of ankyrin was not an artifact of fixing and staining cells. We propose a model in which clustering of neuroglian induces a conformational change in the cytoplasmic domain that drives polarized assembly of the spectrin cytoskeleton.
Jefford
G
G
Department of Neurobiology, Pharmacology, and Physiology, University of Chicago, Chicago, Illinois 60637, USA.
Dubreuil
R R
RR
D000595:Amino Acid Sequence; D000818:Animals; D017487:Ankyrins / Q000096:biosynthesis / Q000378:metabolism*; D001665:Binding Sites; D018801:CD2 Antigens / Q000235:genetics / Q000502:physiology*; D015815:Cell Adhesion Molecules / Q000737:chemistry; D015816:Cell Adhesion Molecules, Neuronal / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D002462:Cell Membrane / Q000502:physiology; D029721:Drosophila Proteins; D004331:Drosophila melanogaster; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000032:analysis; D008969:Molecular Sequence Data; D009414:Nerve Growth Factors / Q000737:chemistry; D051381:Rats; D011956:Receptors, Cell Surface / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D017487:Ankyrins; D018801:CD2 Antigens; D015815:Cell Adhesion Molecules; D015816:Cell Adhesion Molecules, Neuronal; D029721:Drosophila Proteins; D008164:Luminescent Proteins; D009414:Nerve Growth Factors; C504594:Nfasc protein, rat; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; C061745:Nrg protein, Drosophila; D049452:Green Fluorescent Proteins