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Coexpression of ATP-binding cassette proteins ABCG5 and ABCG8 permits their transport to the apical surface

Coexpression of ATP-binding cassette proteins ABCG5 and ABCG8 permits their transport to the apical surface
Coexpression of ATP-binding cassette proteins ABCG5 and ABCG8 permits their transport to the apical surface
Mutations in either ATP-binding cassette (ABC) G5 or ABCG8 cause sitosterolemia, an autosomal recessive disorder of sterol trafficking. To determine the site of action of ABCG5 and ABCG8, we expressed recombinant, epitope-tagged mouse ABCG5 and ABCG8 in cultured cells. Both ABCG5 and ABCG8 underwent N-linked glycosylation. When either protein was expressed individually in cells, the N-linked sugars remained sensitive to Endoglycosidase H (Endo H). When ABCG5 and ABCG8 were coexpressed, the attached sugars were Endo H–resistant and neuraminidase-sensitive, indicating that the proteins were transported to the trans-Golgi complex. The mature, glycosylated forms of ABCG5 and ABCG8 coimmunoprecipitated, consistent with heterodimerization of these two proteins. The Endo H–sensitive forms of ABCG5 and ABCG8 were confined to the endoplasmic reticulum (ER), whereas the mature forms were present in non-ER fractions in cultured hepatocytes. Immunoelectron microscopy revealed ABCG5 and ABCG8 on the plasma membrane of these cells. In polarized WIF-B cells, recombinant ABCG5 localized to the apical (canalicular) membrane when coexpressed with ABCG8, but not when expressed alone. To our knowledge this is the first direct demonstration that trafficking of an ABC half-transporter to the cell surface requires the presence of its dimerization partner.
0021-9738
659-669
Graf, Gregory A.
9997ad6f-9da4-4185-931e-1757e261670d
Li, Wei-Ping
306e2a69-a21e-429e-b75b-27da9465e74e
Gerard, Robert D.
0e10acd8-0ab0-462b-82cd-b5c2542f2e5f
Gelissen, Ingrid
75c4c3dc-bbbb-46e6-b8b1-b9f3e7446a13
White, Ann
495e8275-c090-4f7a-98ca-fa5eb06bd3e9
Cohen, Jonathan C.
1402a549-7e7e-4808-bc2c-5aca8baec701
Hobbs, Helen H.
36279b6a-361d-4c62-a3c1-3ed48071aff3
Graf, Gregory A.
9997ad6f-9da4-4185-931e-1757e261670d
Li, Wei-Ping
306e2a69-a21e-429e-b75b-27da9465e74e
Gerard, Robert D.
0e10acd8-0ab0-462b-82cd-b5c2542f2e5f
Gelissen, Ingrid
75c4c3dc-bbbb-46e6-b8b1-b9f3e7446a13
White, Ann
495e8275-c090-4f7a-98ca-fa5eb06bd3e9
Cohen, Jonathan C.
1402a549-7e7e-4808-bc2c-5aca8baec701
Hobbs, Helen H.
36279b6a-361d-4c62-a3c1-3ed48071aff3

Graf, Gregory A., Li, Wei-Ping, Gerard, Robert D., Gelissen, Ingrid, White, Ann, Cohen, Jonathan C. and Hobbs, Helen H. (2002) Coexpression of ATP-binding cassette proteins ABCG5 and ABCG8 permits their transport to the apical surface. Journal of Clinical Investigation, 110 (5), 659-669. (doi:10.1172/JCI200216000).

Record type: Article

Abstract

Mutations in either ATP-binding cassette (ABC) G5 or ABCG8 cause sitosterolemia, an autosomal recessive disorder of sterol trafficking. To determine the site of action of ABCG5 and ABCG8, we expressed recombinant, epitope-tagged mouse ABCG5 and ABCG8 in cultured cells. Both ABCG5 and ABCG8 underwent N-linked glycosylation. When either protein was expressed individually in cells, the N-linked sugars remained sensitive to Endoglycosidase H (Endo H). When ABCG5 and ABCG8 were coexpressed, the attached sugars were Endo H–resistant and neuraminidase-sensitive, indicating that the proteins were transported to the trans-Golgi complex. The mature, glycosylated forms of ABCG5 and ABCG8 coimmunoprecipitated, consistent with heterodimerization of these two proteins. The Endo H–sensitive forms of ABCG5 and ABCG8 were confined to the endoplasmic reticulum (ER), whereas the mature forms were present in non-ER fractions in cultured hepatocytes. Immunoelectron microscopy revealed ABCG5 and ABCG8 on the plasma membrane of these cells. In polarized WIF-B cells, recombinant ABCG5 localized to the apical (canalicular) membrane when coexpressed with ABCG8, but not when expressed alone. To our knowledge this is the first direct demonstration that trafficking of an ABC half-transporter to the cell surface requires the presence of its dimerization partner.

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Published date: 2002

Identifiers

Local EPrints ID: 26329
URI: http://eprints.soton.ac.uk/id/eprint/26329
ISSN: 0021-9738
PURE UUID: e57615b2-f909-418d-a1a5-1e094199020e

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Date deposited: 20 Apr 2006
Last modified: 15 Jul 2019 19:14

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Contributors

Author: Gregory A. Graf
Author: Wei-Ping Li
Author: Robert D. Gerard
Author: Ingrid Gelissen
Author: Ann White
Author: Jonathan C. Cohen
Author: Helen H. Hobbs

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