Characterization of Oligomeric Human Half-ABC Transporter ATP-binding Cassette G2

Xu, Junkang; Yang, Lei; Yang, Youyun; Bates, Susan E.; Zhang, Jian-Ting

doi:10.1074/jbc.m310785200

Cited by 236 publications

(240 citation statements)

References 33 publications

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“…Higher order oligomeric structures have recently been reported for ABCG2 (30) and ABCA1 (31). Interestingly, ABCA1 appeared to transition from a dimer to a higher order oligomer during the catalytic cycle (31).…”

Section: Discussionmentioning

confidence: 99%

“…Models proposing hydrolysis of one or two ATP molecules per transport cycle have been put forward (26), but measured ratios of ATPase activity to allocrite transport vary widely, even for the same transporter (27)(28)(29)(30). In our previous study using purified recombinant G5G8, sterol-transfer activity was not sufficiently stable over time to allow for a quantitative estimate of the allocrite transfer rate.…”

Section: Discussionmentioning

confidence: 99%

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Purification and Reconstitution of Sterol Transfer by Native Mouse ABCG5 and ABCG8

et al. 2008

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ABCG5 (G5) and ABCG8 (G8) are ATP-binding cassette half-transporters that limit intestinal uptake and promote biliary secretion of neutral sterols. Here, we describe the purification of endogenous G5G8 from mouse liver to near homogeneity. We incorporated the native proteins into membrane vesicles and reconstituted sterol transfer. Native gel electrophoresis, density-gradient ultracentrifugation, and chemical cross-linking studies indicated that the functional native complex is a heterodimer. No higher order oligomeric forms were observed at any stage in the catalytic cycle. Sterol transfer activity by purified native G5G8 was stable, stereospecific, and selective. We also report that G5 but not G8 is S-palmitoylated and that palmitoylation is not essential for dimerization, trafficking, or biliary sterol secretion. Both G5 and G8 have short but highly conserved cytoplasmic tails. The functional roles of these C-terminal regions were examined using an in vivo functional assay. ABCG5 (G5) 1 and ABCG8 (G8) are ABC half-transporters that limit the accumulation of neutral sterols in the body (1). The two proteins are expressed primarily in enterocytes and hepatocytes, where they heterodimerize in the endoplasmic reticulum prior to being transported to apical membranes (1-3). Inactivating mutations in either G5 or G8 cause sitosterolemia, a recessive disorder characterized by hypercholesterolemia, phytosterolemia, and premature coronary artery disease (3,4). The role of G5 and G8 in sterol trafficking in vivo has been † This work was supported by Grant HL72304 from the National Institutes of Health (NIH). NIH Public Access Author ManuscriptBiochemistry. Author manuscript; available in PMC 2009 July 8. Published in final edited form as:Biochemistry. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript examined in detail using genetically modified mice, in which G5 and G8 are either overexpressed or inactivated (5-10). In the intestine, the G5G8 heterodimer limits the absorption of dietary sterols, especially plant-derived sterols (5). G5G8 synthesized in the liver is located in the bile canalicular membrane and is required for efficient secretion of neutral sterols into bile (5).Previously, we developed an in vitro assay using recombinant G5G8 expressed in Sf9 cells to elucidate the mechanism by which G5G8 promotes translocation of sterols across membranes (11). Membrane vesicles prepared from Sf9 cells expressing wild-type G5 and G8 supported the transfer of cholesterol from donor vesicles. G5G8-mediated transfer was stereoselective and specific for neutral sterols. Introduction of mutations predicted to disrupt ATP hydrolysis abolished G5G8-mediated sterol transfer. The recombinant G5G8 transporter was purified to near homogeneity using affinity chromatography, and the purified heterodimer retained ATPdependent sterol transfer activity when incorporated into proteoliposomes (11). These studies provided the first direct evidence that neutral sterols are the primary transport substrate f...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Purification and Reconstitution of Sterol Transfer by Native Mouse ABCG5 and ABCG8

et al. 2008

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“…The ABCG2 protein was discovered only about 15 years ago and is a "half ABC transporter" containing only one TMD consisting of six transmembrane helices and one NBD. The functional form of ABCG2 has been shown to be a homodimer or homo-oligomer (Bhatia, Schafer, & Hrycyna, 2005;Ni, Mark, Cai, & Mao, 2010;Ozvegy et al, 2001;Xu, Liu, Yang, Bates, & Zhang, 2004). Both of these transporters have a "promiscuous" capacity of recognizing a large number of transported substrates.…”

Section: P0065mentioning

confidence: 99%

Lipid Regulation of the ABCB1 and ABCG2 Multidrug Transporters

Hegedüs

Telbisz

Hegedüs

et al. 2015

ABC Transporters and Cancer

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“…HEK293-transfected sublines HEK293/ Vec (18), HEK293/ABCC1 (19), and HEK293/ABCG2 (20) were obtained from Prof. Jian-ting Zhang (Indiana University Simon Cancer Center, Indianapolis, IN). The human breast cancer cell line MCF-7 and the mitoxantrone selected MDR cell line MCF-7/MX (21) were kindly provided by Dr. E. Schneider (Wadsworth Center, Albany, NY).…”

Section: Cell Culture Treatments and Lysate Preparationsmentioning

confidence: 99%

“…Western blot and fluorescence-activated cell sorting (FACS) analyses of drug accumulation were carried out exactly as we previously described (15,20). To determine the conformational change of ABCG2 following treatment with ABCG2 inhibitors, HEK293/ ABCG2 cells were incubated with 0.5 and 2.5 mmol/L sorafenib at 37 C for 10 minutes before PE-conjugated 5D3 antibody (1:20 dilution) was added and incubated for 30 minutes.…”

Section: Western Blot and Fluorescence-activated Cell Sorting Analysesmentioning

confidence: 99%

New Use for an Old Drug: Inhibiting ABCG2 with Sorafenib

Wei

Zhao

et al. 2012

Molecular Cancer Therapeutics

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Human ABCG2, a member of the ATP-binding cassette transporter superfamily, represents a promising target for sensitizing MDR in cancer chemotherapy. Although lots of ABCG2 inhibitors were identified, none of them has been tested clinically, maybe because of several problems such as toxicity or safety and pharmacokinetic uncertainty of compounds with novel chemical structures. One efficient solution is to rediscover new uses for existing drugs with known pharmacokinetics and safety profiles. Here, we found the new use for sorafenib, which has a dual-mode action by inducing ABCG2 degradation in lysosome in addition to inhibiting its function. Previously, we reported some novel dual-acting ABCG2 inhibitors that showed closer similarity to degradation-induced mechanism of action. On the basis of these ABCG2 inhibitors with diverse chemical structures, we developed a pharmacophore model for identifying the critical pharmacophore features necessary for dual-acting ABCG2 inhibitors. Sorafenib forms impressive alignment with the pharmacophore hypothesis, supporting the argument that sorafenib is a potential ABCG2 inhibitor. This is the first article that sorafenib may be a good candidate for chemosensitizing agent targeting ABCG2-mediated MDR. This study may facilitate the rediscovery of new functions of structurally diverse old drugs and provide a more effective and safe way of sensitizing MDR in cancer chemotherapy.

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Characterization of Oligomeric Human Half-ABC Transporter ATP-binding Cassette G2

Cited by 236 publications

References 33 publications

Purification and Reconstitution of Sterol Transfer by Native Mouse ABCG5 and ABCG8

Purification and Reconstitution of Sterol Transfer by Native Mouse ABCG5 and ABCG8

Lipid Regulation of the ABCB1 and ABCG2 Multidrug Transporters

New Use for an Old Drug: Inhibiting ABCG2 with Sorafenib

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