Subcellular targeting and cytoskeletal attachment of SAP97 to the epithelial lateral membrane.
The synapse-associated protein SAP97 is a member of a novel family of cortical cytoskeletal proteins involved in the localization of ion channels at such membrane specializations as synaptic junctions. These multidomain proteins have binding sites for protein 4.1, GKAPs/SAPAPs, voltage- and ligand-gated ion channels and cell-adhesion molecules containing C-terminal T/SXV motifs. In this study, we evaluated the contribution of individual domains in SAP97 to its selective recruitment and attachment to the cortical cytoskeleton in epithelial cells. We find that the PDZ, SH3 and GK domains, as well as the I3 insert in SAP97, are not essential for subcellular targeting, though both PDZ1-2 domains and the I3 insert affect the efficiency of localization. Instead, we show that the first 65 amino acid residues in SAP97, which are absent from SAP90/PSD-95 and SAP102, direct the selective subcellular localization and can mediate at least one point of attachment of SAP97 to the cytoskeleton assembled at sites of cell-cell contact. Our data demonstrate that it is the sequences unique to SAP97 that direct its subcellular targeting to the epithelial lateral membrane.
Department of Neurobiology, University of Alabama at Birmingham, Birmingham, AL 35294-0021, USA.
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Wu
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Reuver
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Kuhlendahl
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Chung
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Garner
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D018938:Caco-2 Cells; D002462:Cell Membrane / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D000074288:Discs Large Homolog 1 Protein; D004847:Epithelial Cells / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006801:Humans; D007473:Ion Channels / Q000378:metabolism; D008164:Luminescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection
D048868:Adaptor Proteins, Signal Transducing; C513314:DLG1 protein, human; D000074288:Discs Large Homolog 1 Protein; D007473:Ion Channels; D008164:Luminescent Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins