ATP-binding cassette transporter A1 contains an NH2-terminal signal anchor sequence that translocates the protein's first hydrophilic domain to the exoplasmic space.
Mutations in the ATP-binding cassette transporter A1 (ABCA1) transporter are associated with Tangier disease and a defect in cellular cholesterol efflux. The amino terminus of the ABCA1 transporter has two putative in-frame translation initiation sites, 60 amino acids apart. A cluster of hydrophobic amino acids form a potentially cleavable signal sequence in this 60-residue extension. We investigated the functional role of this extension and found that it was required for stable protein expression of transporter constructs containing any downstream transmembrane domains. The extension directed transporter translocation across the ER membrane with an orientation that resulted in glycosylation of amino acids immediately distal to the signal sequence. Neither the native signal sequence nor a green fluorescent protein tag, fused at the amino terminus, was cleaved from ABCA1. The green fluorescent protein fusion protein had efflux activity comparable with wild type ABCA1 and demonstrated a predominantly plasma membrane distribution in transfected cells. These data establish a requirement for the upstream 60 amino acids of ABCA1. This region contains an uncleaved signal anchor sequence that positions the amino terminus in a type II orientation leading to the extracellular presentation of an approximately 600-amino acid loop in which loss-of-function mutations cluster in Tangier disease patients.
Fitzgerald
M L
ML
Lipid Metabolism Unit, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.
Mendez
A J
AJ
Moore
K J
KJ
Andersson
L P
LP
Panjeton
H A
HA
Freeman
M W
MW
D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000918:Antibody Specificity; D002784:Cholesterol / Q000378:metabolism; D018076:DNA, Complementary; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D014176:Protein Biosynthesis; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
D018528:ATP-Binding Cassette Transporters; D018076:DNA, Complementary; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D002784:Cholesterol