ABCMdb

ABCG8 p.Gly574Arg

Predicted by SNAP2:	A: D (91%), C: D (95%), D: D (95%), E: D (95%), F: D (95%), H: D (95%), I: D (95%), K: D (95%), L: D (95%), M: D (95%), N: D (95%), P: D (95%), Q: D (95%), R: D (95%), S: D (85%), T: D (95%), V: D (95%), W: D (95%), Y: D (95%),
Predicted by PROVEAN:	A: N, C: D, D: D, E: D, F: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, Y: D,

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[hide] Mutational studies of G553 in TM5 of ABCG2: a resi... Biochemistry. 2006 Apr 25;45(16):5251-60. Polgar O, Ozvegy-Laczka C, Robey RW, Morisaki K, Okada M, Tamaki A, Koblos G, Elkind NB, Ward Y, Dean M, Sarkadi B, Bates SE
Mutational studies of G553 in TM5 of ABCG2: a residue potentially involved in dimerization.
Biochemistry. 2006 Apr 25;45(16):5251-60., 2006-04-25 [PMID:16618113]

Abstract [show]
ABCG2 is an ATP-binding cassette half-transporter conferring resistance to chemotherapeutic agents such as mitoxantrone, irinotecan, and flavopiridol. With its one transmembrane and one ATP-binding domain, ABCG2 is thought to homodimerize for function. One conserved region potentially involved in dimerization is a three-amino acid sequence in transmembrane segment 5 (residues 552-554). Mutations in the corresponding residues in the Drosophila white protein (an orthologue of ABCG2) are thought to disrupt heterodimerization. We substituted glycine 553 with leucine (G553L) followed by stable transfection in HEK 293 cells. The mutant was not detectable on the cell surface, and markedly reduced protein expression levels were observed by immunoblotting. A deficiency in N-linked glycosylation was suggested by a reduction in molecular mass compared to that of the 72 kDa wild-type ABCG2. Similar results were observed with the G553E mutant. Confocal microscopy demonstrated mostly ER localization of the G553L mutant in HEK 293 cells, even when coexpressed with the wild-type protein. Despite its altered localization, the G553L and G553E mutants were cross-linked using amine-reactive cross-linkers with multiple arm lengths, suggesting that the monomers are in the proximity of each other but are unable to complete normal trafficking. Interestingly, when expressed in Sf9 insect cells, G553L moves to the cell membrane but is unable to hydrolyze ATP or transport the Hoechst dye. Still, when coexpressed, the mutant interferes with the Hoechst transport activity of the wild-type protein. These data show that glycine 553 is important for protein trafficking and are consistent with, but do not yet prove, its involvement in ABCG2 homodimerization.

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231 On the other hand, the ABCG8 G574R mutant, although having some effect on ABCG8 maturation, did not prevent heterodimerization (33). Login to comment

[hide] Genetic basis of sitosterolemia. Curr Opin Lipidol. 2001 Apr;12(2):141-9. Lee MH, Lu K, Patel SB
Genetic basis of sitosterolemia.
Curr Opin Lipidol. 2001 Apr;12(2):141-9., [PMID:11264985]

Abstract [show]
The molecular mechanisms regulating the amount of dietary cholesterol retained by the body, as well as the body's ability to exclude other dietary sterols selectively, are poorly understood. An average Western diet will contain approximately 250-500 mg of dietary cholesterol and approximately 200-400 mg of non-cholesterol sterols, of which plant sterols are the major constituents. Approximately 50-60% of dietary cholesterol is absorbed and retained by the normal human body, but less than 1% of the non-cholesterol sterols are retained. There thus exists a subtle mechanism that allows the body to distinguish between cholesterol and non-cholesterol sterols. In sitosterolemia, a rare autosomal recessive disorder, affected individuals hyperabsorb and retain not only cholesterol but also all other sterols, including plant and shellfish sterols from the intestine. Consequently, patients with this disease have very high levels of plant sterols in the plasma, and develop tendon and tuberous xanthomas, accelerated atherosclerosis, and premature coronary artery disease. The STSL locus has been mapped to human chromosome 2p21. Mutations in two tandem ABC genes, ABCG5 and ABCG8, encoding sterolin-1 and -2, respectively, are now known to be mutant in sitosterolemia. The identification of these genes should now lead to a better understanding of the molecular mechanism(s) governing the highly selective absorption and retention of cholesterol by the body. Indeed, it is the very existence of this disease that has given credence to the hypothesis that there is a molecular pathway that regulates dietary cholesterol absorption and sterol excretion by the body.

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108 Mutations in ABCG5 and ABCG8 in sitosterolemia Patients Nationality/ethnicity ABCG5 ABCG8 4 US/Caucasian Arg412X Trp361X 9 US/Caucasian Arg543Ser Gln172X 56* US/Caucasian Tyr658X Trp361X 60 US/Caucasian Trp361X ± 90* US/Caucasian Trp361X Trp361X 94 US/Caucasian Trp361X Arg184His 120 US/Caucasian Leu501Pro Trp361X 125 US/Caucasian Trp361X Trp361X 128 US/Caucasian Leu596Arg ± 32 SA Caucasian Arg121X Arg121X 98 Dutch Caucasian Gly574Glu Trp361X 102* US/Caucasian Trp361X ± 135 Columbian/Caucasian Trp536X Trp536X 20 Finnish Trp361X Trp361X 116 Norwegian Trp36X Trp361X 84* US/Amish/Mennonite Gly574Arg Gly574Arg 108 US/Amish/Mennonite Gly574Arg Gly574Arg 25 SA/Asian Arg243X Arg243X 40 Japanese Arg419His Arg419His 46 Japanese Arg389His Arg389His 63 Japanese del Exon 3 del Exon 3 113 Japanese Arg389His Arg389His 132 Japanese Arg419His ± 140 Japanese Arg408X Arg408X 146 Japanese Arg389His Arg389His 157 US/Caucasian Arg419Pro Arg419Pro 15 US/Caucasian ± ± 143 SA/Indian Asian ± ± Arg121X Arg164X 149 African American Glu145Gln ± 1* German/Swiss Trp361X Trp361X 2* US/Amish Gly574Arg Gly574Arg 3* US/Caucasian Trp361X Tyr658X 5* US/Caucasian Trp361X Arg412X 6* US/Caucasian Leu596Arg ± 7 US/Hispanic Arg412X del547C4191X 8* NZ/Caucasian Trp361X ± 4 Chinese Arg263Gln Pro231Thr 9 Chinese Arg408X ± This is a compilation of the mutations identified in ABCG5 and ABCG8. Login to comment

[hide] Mutations in the human ATP-binding cassette transp... Hum Mutat. 2002 Aug;20(2):151. Heimerl S, Langmann T, Moehle C, Mauerer R, Dean M, Beil FU, von Bergmann K, Schmitz G
Mutations in the human ATP-binding cassette transporters ABCG5 and ABCG8 in sitosterolemia.
Hum Mutat. 2002 Aug;20(2):151., [PMID:12124998]

Abstract [show]
Phytosterolemia or Sitosterolemia is a rare autosomal recessive disorder characterized by highly elevated plasma levels of plant sterols and cholesterol as a consequence of hyperabsorption and impaired biliary secretion of sterols. The disease is caused by mutations in two half size ATP-binding cassette transporters, ABCG5 and ABCG8. We have analyzed the genomic sequence of ABCG5 and ABCG8 in five well-characterized patients with Sitosterolemia. In the first patient we found a heterozygous mutation in exon 8 of the ABCG5 gene leading to a premature termination of the protein (Arg408Ter). This German patient is the first European showing a mutation of the ABCG5 gene. In a second patient we found a novel heterozygous mutation in exon 5 of ABCG8 (c.584T>A; Leu195Gln). Both patients were heterozygous for the identified mutation, but no mutation could be identified on the other chromosome. In three further analyzed patients we found mutations in exons 7, 9 and 11 of the ABCG8 gene, respectively, of which two result in a premature termination signal for translation products. One of these patients was compound heterozygous (Trp361Ter and Arg412Ter), the other was homozygous for Trp361Ter. The third patient was homozygous for an amino acid exchange (Gly574Arg). In conclusion this report describes one novel mutation affecting a highly conserved amino acid and two previously identified mutations in the ABCG8 gene. In addition, we identified for the first time a mutation in the ABCG5 gene of a European Sitosterolemia patient.

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No. Sentence Comment
77 C Y658stop R121stop R164stop Q172stop R184H L195Q P231T G574R G574E L572P L596R N ABC B S AA R263Q E146Q R405H R543S W536stop R412stop W361stop C R419P R419H R408stop R398H N437K R550S R243stop N ABCG5 ABCG8 S B A IVS1 -2A>G Del547C>191stop L501P L596R 1568_1572delTCTTT 1798_1800delTTC Del Exon 3 C336-337insA 201 * Signature 250 ABCG1 Q..EKDEG.R REMVKEILTA L GLLSCANTR TGS.... .LS GGQR KRLAIA ABCG2 ATTMTNHE.K NERINRVIEE L GLDKVADSK VGTQFIR GVS GGER KRTSIG ABCG4 S..EKQEV.K KELVTEILTA L GLMSCSHTR TAL.... .LS GGQR KRLAIA ABCG5 R..RGNPGSF QKKVEAVMAE L SLSHVADRL IGNYSLG GIS TGER RRVSIA ABCG8 PRTFSQAQ.R DKRVEDVIAE L RLRQCADTR VGNMYVR GLS GGER RRVSIG Figure 2: Alignment of the human ABC transporters G1, G2, G4, G5 and G8. The amino acid change Leu195Gln in ABCG8 found in patient 2 is located intracellularly between the Walker A and the Signature C-motif. Login to comment
12 The third patient was homozygous for an amino acid exchange (Gly574Arg). Login to comment
37 (mg/dl) Gene Mutation Allele 1 Mutation Allele 2 Ethnicity 1 305 246 38 101 31.1 ABCG5 Arg408X - Caucasian / German 2 210 129 67 72 20.3 ABCG8 Leu195Gln - Caucasian 3 218 166 35 84 22.1 ABCG8 Trp361X Arg412X Caucasian 4 247 155 56 189 20.5 ABCG8 Trp361X Trp361X Caucasian 5 214 145 42 90 17.5 ABCG8 Gly574Arg Gly574Arg Caucasian / Swiss Patient 1 was a man who suffered from a myocardial infarction at the age of 31 years. Login to comment
48 Patient 5 carried a homozygous mutation in exon 11(c.1720 G>A) of the ABCG8 gene resulting in an amino acid exchange at position 574 (Gly574Arg). Login to comment
80 Patient 3 carried a compound heterozygous mutation resulting in a truncated protein (Trp361X and Arg412X), whereas patient 4 and patient 5 had homozygous mutations for Trp361X and Gly574Arg, respectively. Login to comment

[hide] Missense mutations in ABCG5 and ABCG8 disrupt hete... J Biol Chem. 2004 Jun 4;279(23):24881-8. Epub 2004 Mar 30. Graf GA, Cohen JC, Hobbs HH
Missense mutations in ABCG5 and ABCG8 disrupt heterodimerization and trafficking.
J Biol Chem. 2004 Jun 4;279(23):24881-8. Epub 2004 Mar 30., 2004-06-04 [PMID:15054092]

Abstract [show]
Mutations in ABCG5 (G5) or ABCG8 (G8) cause sitosterolemia, an autosomal recessive disease characterized by sterol accumulation and premature atherosclerosis. G5 and G8 are ATP-binding cassette (ABC) half-transporters that must heterodimerize to move to the apical surface of cells. We examined the role of N-linked glycans in the formation of the G5/G8 heterodimer to gain insight into the determinants of folding and trafficking of these proteins. Site-directed mutagenesis revealed that two asparagine residues (Asn(585) and Asn(592)) are glycosylated in G5 and that G8 has a single N-linked glycan attached to Asn(619). N-Linked glycosylation of G8 was required for efficient trafficking of the G5/G8 heterodimer, but mutations that abolished glycosylation of G5 did not prevent trafficking of the heterodimer. Both G5 and G8 are bound by the lectin chaperone, calnexin, suggesting that the calnexin cycle may facilitate folding of the G5/G8 heterodimer. To determine the effects of 13 disease-causing missense mutations in G5 and G8 on formation and trafficking of the G5/G8 heterodimer, mutant forms of the half-transporters were expressed in CHO-K1 cells. All 13 mutations reduced trafficking of the G5/G8 heterodimer from the endoplasmic reticulum to the Golgi complex, and most prevented the formation of stable heterodimers between G5 and G8. We conclude that the majority of the molecular defects in G5 and G8 that cause sitosterolemia impair transport of the sterol transporter to the cell surface.

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198 Three exceptions, L596R, G574E, and G574R in G8, result in the substitution of charged for neutral amino acids. Login to comment
206 The remaining two mutations (R543S and G574R) decreased, but did not eliminate, the proportion of G8 present in the mature form. Login to comment
219 Only those mutant forms of G8 that were trafficking-competent (R543S and G574R) and to a lesser extent L596R were detected in the position of the dimer. Login to comment
197 Three exceptions, L596R, G574E, and G574R in G8, result in the substitution of charged for neutral amino acids. Login to comment
205 The remaining two mutations (R543S and G574R) decreased, but did not eliminate, the proportion of G8 present in the mature form. Login to comment
218 Only those mutant forms of G8 that were trafficking-competent (R543S and G574R) and to a lesser extent L596R were detected in the position of the dimer. Login to comment

[hide] Phytosterols and phytosterolemia: gene-diet intera... Genes Nutr. 2011 Feb;6(1):17-26. Epub 2010 Aug 28. Izar MC, Tegani DM, Kasmas SH, Fonseca FA
Phytosterols and phytosterolemia: gene-diet interactions.
Genes Nutr. 2011 Feb;6(1):17-26. Epub 2010 Aug 28., [PMID:21437027]

Abstract [show]
Phytosterol intake is recommended as an adjunctive therapy for hypercholesterolemia, and plant sterols/stanols can reduce cholesterol absorption at the intestinal lumen through the Niemann-Pick C1 Like 1 (NPC1L1) transporter pathway by competitive solubilization in mixed micelles. Phytosterol absorption is of less magnitude than cholesterol and is preferably secreted in the intestinal lumen by ABCG5/G8 transporters. Therefore, plasma levels of plant sterols/stanols are negligible compared with cholesterol, under an ordinary diet. The mechanisms of cholesterol and plant sterols absorption and the whole-body pool of sterols are discussed in this chapter. There is controversy about treatment with statins inducing further increase in plasma non-cholesterol sterols raising concerns about the safety of supplementation of plant sterols to such drugs. In addition, increase in plant sterols has also been reported upon consumption of plant sterol-enriched foods, regardless of other treatments. Rare mutations on ABCG5/G8 transporters affecting cholesterol/non-cholesterol extrusion, causing sitosterolemia with xanthomas and premature atheroslerotic disease are now known, and cholesterol/plant sterols absorption inhibitor, ezetimibe, emerges as the drug that reduces phytosterolemia and promotes xanthoma regression. On the other hand, common polymorphisms affecting the NPC1L1 transporter can interfere with the action of ezetimibe. Gene-diet interactions participate in this intricate network modulating the expression of genetic variants on specific phenotypes and can also affect the individual response to the hypolipidemic treatment. These very interesting aspects promoted a great deal of research in the field.

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111 The presence of the homozygous mutation Gly574Arg in the ABCG8 gene has been found previously in the Amish-Mennonite patients where a founder effect has been reported [73]. Login to comment
113 The presence of the homozygous mutation Gly574Arg in the ABCG8 gene has been found previously in the Amish-Mennonite patients where a founder effect has been reported [73]. Login to comment

[hide] Sitosterolaemia in Switzerland: molecular genetics... Clin Genet. 2005 Aug;68(2):174-8. Solca C, Stanga Z, Pandit B, Diem P, Greeve J, Patel SB
Sitosterolaemia in Switzerland: molecular genetics links the US Amish-Mennonites to their European roots.
Clin Genet. 2005 Aug;68(2):174-8., [PMID:15996216]

Abstract [show]
Sitosterolaemia is a rare autosomal recessive disease characterized by increased intestinal absorption of plant sterols, decreased hepatic excretion into bile and elevated concentrations in plasma phytosterols. Homozygous or compound heterozygous loss of function mutations in either of the ATP-binding cassette (ABC) proteins ABCG5 and ABCG8 explain the increased absorption of plant sterols. Here we report a Swiss index patient with sitosterolaemia, who presented with the classical symptoms of xanthomas, but also had mitral and aortic valvular heart disease. Her management over the last 20 years included a novel therapeutic approach of high-dose cholesterol feeding that was semi-effective. Mutational and extended haplotype analyses showed that our patient shared this haplotype with that of the Amish-Mennonite sitosterolaemia patients, indicating they are related ancestrally.

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56 Mutational analysis showed that our patient was homozygous for the Gly574Arg mutation in ABCG8. Login to comment
64 These data support the contention that the Gly574Arg mutation in the Amish-Mennonite originated in Europe and is likely more than 250 years old. Login to comment

[hide] Genetic disorders associated with ATP binding cass... Mol Genet Metab. 2002 Sep-Oct;77(1-2):13-20. Burris TP, Eacho PI, Cao G
Genetic disorders associated with ATP binding cassette cholesterol transporters.
Mol Genet Metab. 2002 Sep-Oct;77(1-2):13-20., [PMID:12359125]

Abstract [show]
Coronary artery disease is the most prevalent form of mortality and morbidity in Western countries. Studies in the last several decades have identified high LDL cholesterol and low HDL cholesterol as major risk factors leading to the disease. Human genetic studies have provided significant insight into the regulation of lipoprotein metabolism. In the last several years, the genes associated with several rare genetic diseases of lipid metabolism have been revealed. These landmark discoveries that identified mutant ABC cholesterol transporters as the underlying causes of these genetic disorders have paved the way for better understanding of the cellular cholesterol transport process and HDL biogenesis. This summary provides an overview and discussion of the most recent progress that includes molecular mechanism and regulation of cholesterol transport mediated by these ABC transporters.

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112 Regulation of ABC cholesterol transporters by LXR/ RXR heterodimer One feature of ABC transporters is that they are usually feed back/forward regulated by their own sub-Table 2 Summary of mutations described in sitosterolemia Nucleotide sequence change Protein sequence change ABC transporters References 402Del Truncated protein ABCG5 [12] C867T R243X ABCG5 [12] G1306A R389H ABCG5 [12] C1362T R408X ABCG5 [11,12] G1396A R419H ABCG5 [12] G1396C R419P ABCG5 [12] 547Del P231T ABCG8 [11] A691C 191Stop ABCG8 [11] G788A R263Q ABCG8 [11] G1083A W361Stop ABCG8 [11] C1234T R412stop ABCG8 [11] G1720A G574R ABCG8 [11] T1787G L596R ABCG8 [11] C1974G Y658Stop ABCG8 [11] strates at the level of transcription. Login to comment

[hide] ATP binding cassette transporter G1 (ABCG1) is an ... Proc Natl Acad Sci U S A. 2011 Dec 6;108(49):19719-24. Epub 2011 Nov 17. Tarling EJ, Edwards PA
ATP binding cassette transporter G1 (ABCG1) is an intracellular sterol transporter.
Proc Natl Acad Sci U S A. 2011 Dec 6;108(49):19719-24. Epub 2011 Nov 17., [PMID:22095132]

Abstract [show]
Four members of the mammalian ATP binding cassette (ABC) transporter G subfamily are thought to be involved in transmembrane (TM) transport of sterols. The residues responsible for this transport are unknown. The mechanism of action of ABCG1 is controversial and it has been proposed to act at the plasma membrane to facilitate the efflux of cellular sterols to exogenous high-density lipoprotein (HDL). Here we show that ABCG1 function is dependent on localization to intracellular endosomes. Importantly, localization to the endosome pathway distinguishes ABCG1 and/or ABCG4 from all other mammalian members of this superfamily, including other sterol transporters. We have identified critical residues within the TM domains of ABCG1 that are both essential for sterol transport and conserved in some other members of the ABCG subfamily and/or the insulin-induced gene 2 (INSIG-2). Our conclusions are based on studies in which (i) biotinylation of peritoneal macrophages showed that endogenous ABCG1 is intracellular and undetectable at the cell surface, (ii) a chimeric protein containing the TM of ABCG1 and the cytoplasmic domains of the nonsterol transporter ABCG2 is both targeted to endosomes and functional, and (iii) ABCG1 colocalizes with multiple proteins that mark late endosomes and recycling endosomes. Mutagenesis studies identify critical residues in the TM domains that are important for ABCG1 to alter sterol efflux, induce sterol regulatory element binding protein-2 (SREBP-2) processing, and selectively attenuate the oxysterol-mediated repression of SREBP-2 processing. Our data demonstrate that ABCG1 is an intracellular sterol transporter that localizes to endocytic vesicles to facilitate the redistribution of specific intracellular sterols away from the endoplasmic reticulum (ER).

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164 Interestingly, a point mutation in this residue in ABCG8 (G574R) has been identified in a patient with sitosterolemia (38). Login to comment

[hide] Two genes that map to the STSL locus cause sitoste... Am J Hum Genet. 2001 Aug;69(2):278-90. Epub 2001 Jul 9. Lu K, Lee MH, Hazard S, Brooks-Wilson A, Hidaka H, Kojima H, Ose L, Stalenhoef AF, Mietinnen T, Bjorkhem I, Bruckert E, Pandya A, Brewer HB Jr, Salen G, Dean M, Srivastava A, Patel SB
Two genes that map to the STSL locus cause sitosterolemia: genomic structure and spectrum of mutations involving sterolin-1 and sterolin-2, encoded by ABCG5 and ABCG8, respectively.
Am J Hum Genet. 2001 Aug;69(2):278-90. Epub 2001 Jul 9., [PMID:11452359]

Abstract [show]
Sitosterolemia is a rare autosomal recessive disorder characterized by (a) intestinal hyperabsorption of all sterols, including cholesterol and plant and shellfish sterols, and (b) impaired ability to excrete sterols into bile. Patients with this disease have expanded body pools of cholesterol and very elevated plasma plant-sterol species and frequently develop tendon and tuberous xanthomas, accelerated atherosclerosis, and premature coronary artery disease. In previous studies, we have mapped the STSL locus to human chromosome 2p21. Recently, we reported that a novel member of the ABC-transporter family, named "sterolin-1" and encoded by ABCG5, is mutated in 9 unrelated families with sitosterolemia; in the remaining 25 families, no mutations in sterolin-1 could be identified. We identified another ABC transporter, located <400 bp upstream of sterolin-1, in the opposite orientation. Mutational analyses revealed that this highly homologous protein, termed "sterolin-2" and encoded by ABCG8, is mutated in the remaining pedigrees. Thus, two highly homologous genes, located in a head-to-head configuration on chromosome 2p21, are involved as causes of sitosterolemia. These studies indicate that both sterolin-1 and sterolin-2 are indispensable for the regulation of sterol absorption and excretion. Identification of sterolin-1 and sterolin-2 as critical players in the regulation of dietary-sterol absorption and excretion identifies a new pathway of sterol transport.

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134 A human full- Table 2 Compilation of Mutations in ABCG5 and ABCG8 PATIENT (NATIONALITY/ETHNICITY) MUTATIONS IN ABCG5a ABCG8b 4 (U.S./white) Trp361X (1173GrA) / Arg412X (1324CrT) 9 (U.S./white) Arg543Ser (1719GrT) / Gln172X (604CrT) 56c (U.S./white) Trp361X (1173GrA) / Tyr658X (2064CrG) 60 (U.S./white) Trp361X (1173GrA) / IVS1 -2 ArG 90c (U.S./white) Trp361X (1173GrA) / Trp361X (1173GrA) 94 (U.S./white) Trp361X (1173GrA) / Arg184His (641GrA) 120 (U.S./white) Trp361X (1173GrA) / Leu501Pro (1592TrC) 125 (U.S./white) Trp361X (1173GrA) / Trp361X (1173GrA) 128 (U.S./white) Leu596Arg (1877TrG) / IVS1 -2 ArG 172 (U.S./white) Trp361X (1173GrA) / Tyr658Stop (2064CrG) 166c (U.S./white) Trp361X (1173GrA) / Arg412X (1324CrT) 32 (SA/white) Arg121X (451CrT) / Arg121X (451CrT) 98 (Dutch/white) Trp361X (1173GrA) / Gly574Glu (1811GrA) 102c,d (U.S./white) Trp361X (1173GrA) / … 84c (U.S./Amish-Mennonite) Gly574Arg (1810GrA) / Gly574Arg (1810GrA) 108c (U.S./Amish-Mennonite) Gly574Arg (1810GrA) / Gly574Arg (1810GrA) 135 (Columbian/white) Trp536X (1698GrA) / Trp536X (1698GrA) 175 (French) 1798_1800delTTC / Arg405His (1304GrA) 20 (Finnish) Trp361X (1173GrA) / Trp361X (1173GrA) 154 (Finnish) Trp361X (1173GrA) / … 116 (Norwegian) Trp361X (1173GrA) / Trp361X (1173GrA) 163 (Swedish) Trp361X (1173GrA) / Leu572Pro (1805TrC) 15 (U.S./white) 1568_1572delTCTTT / IVS1 -2 ArG 143 (SA/Asian) Arg164X (580CrT) / Arg121X (451CrT) 25 (SA/Asian) Arg243X (876CrT) / Arg243X (876CrT) 40 (Japanese) Arg419His (1396GrA) / Arg419His (1396GrA) 46 (Japanese) Arg389His (1306GrA) / Arg389His (1306GrA) 63 (Japanese) del exon 3 / del exon 3 113 (Japanese) Arg389His (1306GrA) / Arg389His (1306GrA) 132 (Japanese) Arg419His (1396GrC) / Arg550Ser (1790ArC) 140 (Japanese) Arg408X (1362CrT) / Arg408X (1362CrT) 146 (Japanese) Arg389His (1306GrA) / Arg389His (1306GrA) 157 (U.S./white) Arg419Pro (1396GrC) / Arg419Pro (1396GrC) 149 (African American) Glu146Gln (576GrC) / … 1c,e (German/Swiss) Trp361X / Trp361X 2c,e (U.S./Amish) Gly574Arg / Gly574Arg 3c,e (U.S./white) Trp361X / Tyr658X 5c,d (U.S./white) Trp361X / Arg412X 6c,e (U.S./white) Leu596Arg / … 7d (U.S./Hispanic) Arg412X / del547Cr191X 8c,e (New Zealand/white) Trp361X / … 4e (Chinese) Arg263Gln / Pro231Thr 9e (Chinese) Arg408X / … a GenBank accession number AF312715. Login to comment
146 of Alleles Frequency Restriction-Enzyme Recognition ABCG5: Glu146Gln 1 .05 Gain of AlwNI Arg243X 2 .10 Gain of AlwNI Arg389His 6 .30 Loss of BstUI Arg408X 3 .15 Loss of AvaI Arg419Pro 2 .10 Loss of BstUI Arg419His 3 .15 Loss of BstUI del exon 3 2 .10 … Arg550Ser 1 .05 … Total 20 ABCG8: Arg121X 3 .061 Gain of DdeI Arg164stop 1 .020 … Gln172X 1 .020 Gain of BfaI Arg184His 1 .020 Gain of NalIII Pro231Thr 1 .020 Loss of NlaIV Arg263Gln 1 .020 Gain of AluI Trp361X 19 .39 … Arg405His 1 .020 … Arg412X 3 .061 Gain of DdeI Leu501Pro 1 .020 Loss of AluI Trp536X 2 .041 Gain of AhdI Arg543Ser 1 .020 … Leu572Pro 1 .020 Gain of FauI Gly574Glu 1 .020 Loss of MspI Gly574Arg 4 .082 Loss of MspI Leu596Arg 1 .020 Gain of MspI Tyr658X 2 .041 Gain of SfcI IVS1 -2ArG 3 .061 Gain of BtgI 1798_1800delTTC 1 .020 … 1568_1572delTCTTT 1 .020 … Total 49 Mutations of Sterolin-2/ABCG8 as the Cause of Sitosterolemia Information on the exon/intron boundaries was used to screen probands, including those known to be mutated for sterolin-1, and to compare them to normal controls. Login to comment

[hide] The ABCG8 G574R variant, serum plant sterol levels... Arterioscler Thromb Vasc Biol. 2013 Feb;33(2):413-9. doi: 10.1161/ATVBAHA.112.245480. Epub 2012 Dec 13. Horenstein RB, Mitchell BD, Post WS, Lutjohann D, von Bergmann K, Ryan KA, Terrin M, Shuldiner AR, Steinle NI
The ABCG8 G574R variant, serum plant sterol levels, and cardiovascular disease risk in the Old Order Amish.
Arterioscler Thromb Vasc Biol. 2013 Feb;33(2):413-9. doi: 10.1161/ATVBAHA.112.245480. Epub 2012 Dec 13., [PMID:23241408]

Abstract [show]
OBJECTIVE: To determine whether long-term exposure to moderate elevations in plasma plant sterol levels increases risk for atherosclerosis. METHODS AND RESULTS: In Old Order Amish participants aged 18 to 85 years, with (n=110) and without (n=181) 1 copy of the ABCG8 G574R variant, we compared mean plasma levels of plant sterols and cholesterol precursors and carotid intima-media wall thickness. Carriers of a single 574R allele had increased plant sterol levels (eg, 35%-37% higher plasma levels of sitosterol, campesterol, and stigmasterol) and increased plant sterol/cholesterol ratios (P<0.001 for all). 574R carriers had significantly decreased levels of lathosterol and lanosterol, precursors in a pathway for endogenous cholesterol synthesis, suggesting that plant sterols may alter regulation of genes involved in cholesterol synthesis. The G574R variant was not associated with high-density lipoprotein cholesterol or low-density lipoprotein cholesterol levels. Compared with noncarriers, 574R carriers had decreased carotid intima-media wall thickness (0.62 versus 0.66 mm; age- and sex-adjusted P=0.03). Adjustment for body weight, blood pressure, and standard lipid measures (high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, and triglycerides) did not alter this association. CONCLUSIONS: Although the G574R variant is associated with moderately elevated plant sterol levels, carriers of the 574R allele had modestly lower levels of carotid wall thickness compared with noncarriers.

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13 Methods and Results-In Old Order Amish participants aged 18 to 85 years, with (n=110) and without (n=181) 1 copy of the ABCG8 G574R variant, we compared mean plasma levels of plant sterols and cholesterol precursors and carotid intima-media wall thickness. Login to comment
16 The G574R variant was not associated with high-density lipoprotein cholesterol or low-density lipoprotein cholesterol levels. Login to comment
19 Conclusion-Although the G574R variant is associated with moderately elevated plant sterol levels, carriers of the 574R allele had modestly lower levels of carotid wall thickness compared with noncarriers.ߒ Èa;Èa;(Arterioscler Thromb Vasc Biol. Login to comment
21 Key Words: ABC transporter fc; atherosclerosis fc; lipids fc; plant sterols fc; sitosterol The ABCG8 G574R Variant, Serum Plant Sterol Levels, and Cardiovascular Disease Risk in the Old Order Amish Richard B. Horenstein, Braxton D. Mitchell, Wendy S. Post, Dieter L&#fc;tjohann, Klaus von Bergmann, Kathleen A. Ryan, Michael Terrin, Alan R. Shuldiner, Nanette I. Steinle Simvastatin Survival Study, individuals within the upper quartile of campesterol:cholesterol ratios were more likely to require statin titration and less likely to receive risk reduction from simvastatin.8,9 It has also been reported that postmenopausal women with heart disease had higher phytosterol concentrations than their respective controls.10 In the Prospective Cardiovascular M&#fc;nster Study, sitosterol concentrations were associated with increased risk of major coronary events in men at high risk of coronary heart disease.11 By contrast, Silbernagel found no association between increased absorption of plant sterol levels and coronary disease,12 nor were elevated levels of plant sterol associated with atherosclerosis in middle-aged men and women in the Dallas Heart Study.13 The potential role of plant sterols in CVD pathogenesis has become a topic of clinical interest with the finding that long-term high dose HMG-CoA reductase inhibitor (statin) use increases plasma plant sterol levels,9 and that ezetimibe, a drug used to block intestinal absorption of sterols, lowers plasma plant sterol levels.14 The prevalent use of plant sterol-enriched foods (eg, stanol margarines) to lower low-density lipoprotein cholesterol (LDL-C) levels adds to the interest in the relation of phytosterols and CVD risk. Login to comment
22 Sitosterolemia was initially identified in 2 sisters of Amish-Mennonite background,15 and the disease was further characterized in additional members of the Amish, including in a 13-year-old boy from the Old Order Amish community in Lancaster County, Pennsylvania who died from coronary artery disease.16 The disease in the boy resulted from a single-nucleotide change, at position 574 in ABCG8, leading to an amino acid substitution from glycine to arginine (Gly574Arg, or G574R; rs137852988). Login to comment
23 Because of the unique ancestral background of the Lancaster County Amish, we hypothesized that this community would have multiple members who are heterozygous for 1 copy of the ABCG8 mutation, and that these individuals would demonstrate moderate elevations in plasma plant sterols and mildly accelerated atherosclerosis. We sought to identify individuals heterozygous for the ABCG8 G574R mutation to further study the role of plant sterols in CVD and to evaluate the association of the carrier or heterozygous status, with subclinical atherosclerosis as assessed by ultrasound measurement of carotid artery intima-media wall thickness (IMT). Login to comment
24 Methods Subjects and Study Design We screened banked DNA samples from 984 participants from our ongoing studies of complex genetic diseases and traits in the Lancaster County Amish17-19 to identify carriers for the ABCG8 G574R mutation and then recruited family members of these individuals to increase the yield of carriers. Login to comment
57 We then recruited 290 family members of these carriers of whom 99 were carriers, thereby increasing our yield to 114 G574R heterozygote carriers, 190 noncarriers, and 1 574R homozygote. Login to comment
62 Thus the final sample included 110 G574R heterozygote carriers and 181 noncarriers. Login to comment
81 Table 1.ߓ Mean Lipid Levels (&#b1;SE) and Baseline Characteristics of Old Order Amish ABCG8 G574R Variant Heterozygotes and Noncarriers (G574G) G574R Heterozygotes (n=110) G574G (n=181) Age, Sex, and BMI-Adjusted Pߙߙ* Age, y 42.2&#b1;2.0 47.2&#b1;1.7 0.01 Sex, % male 45.5 51.9 0.14 BMI, kg/m2 25.8&#b1;0.6 26.5&#b1;0.5 0.22 Total cholesterol, mg/dL 212&#b1;5.8 217&#b1;5.7 0.40 HDL-cholesterol, mg/dL 49.7&#b1;1.7 51.2&#b1;1.5 0.39 LDL-cholesterol, mg/dL 148&#b1;5.4 149&#b1;5.2 0.89 Glucose, mg/dL 87.6&#b1;2.5 90.2&#b1;2.1 0.30 Systolic blood pressure, mmߕHg 123&#b1;1.9 124&#b1;1.5 0.54 Diastolic blood pressure, mmߕHg 75&#b1;1.1 76&#b1;0.9 0.18 Type 2 diabetes mellitus, % 1.8 3.3 >0.95 History of hypertension, % 5.7 14.4 0.17 History of heart surgery, % 0.0 2.3 0.19 History of MI, % 0.0 1.7 0.17 BMI indicates body mass index; HDL, high-density lipoprotein; LDL, low-density lipoprotein; and MI, myocardial infarction. Login to comment
83 Because cIMT is strongly associated with aging, we assessed whether the association of the G574R variant with IMT was modified by age by including a single-nucleotide polymorphism &#d7; age interaction term in the model. Login to comment
85 Discussion The Old Order Amish provide a unique opportunity to test the hypothesis that modestly elevated plant sterol levels are associated with increased subclinical atherosclerosis as the ABCG8 G574R mutation is enriched in this population and we were able to identify additional carriers through targeted recruitment of families. Login to comment
88 The ABCG8 G574R variant present in the Lancaster Amish has been found in contemporary Switzerland.23 Haplotype analyses have revealed that the initial Amish index cases and the contemporary case identified in Switzerland share a common haplotype, consistent with a common ancestral source for this variant that originated in Switzerland (or before) and was carried to Pennsylvania by ࣙ1 emigrants in the early 1700s. Login to comment
93 As our major conclusion from this study, we find no evidence to Table 2.ߓ Mean Levels (&#b1;SE) of Plant Sterols, Cholesterol Precursors, Sterol/Total Cholesterol Ratios in Old Order Amish ABCG8 G574R Variant Heterozygotes (G574R) and Noncarriers (G574G) G574R Heterozygotes (n=110) G574G (n=181) Age-and Sex-Adjusted P Plant sterols ߓ Sitosterol, mg/dL 0.47&#b1;0.02 0.34&#b1;0.02 <0.0001 ߓ Sitosterol/chol ratio* 2.13&#b1;0.07 1.52&#b1;0.06 <0.0001 ߓ Campesterol, mg/dL 0.60&#b1;0.03 0.44&#b1;0.03 <0.0001 ߓ Campesterol/chol ratio 2.72&#b1;0.10 1.97&#b1;0.08 <0.0001 ߓStigmasterol, bc;g/dL 16.29&#b1;0.84 12.06&#b1;0.79 <0.0001 ߓ Stigmasterol/chol ratio 0.07&#b1;0.003 0.05&#b1;0.003 <0.0001 ߓAvenasterol 1.65&#b1;0.09 1.20&#b1;0.08 <0.0001 ߓ Avena/chol ratio 7.47&#b1;0.40 5.43&#b1;0.35 <0.0001 ߓBrassicasterol, bc;g/dL 26.27&#b1;1.89 22.08&#b1;1.71 0.03 ߓ Brassicasterol/chol ratio 0.12&#b1;0.008 0.010&#b1;0.006 0.008 Proxy for cholesterol absorption ߓ Cholestanol, mg/dL 0.38&#b1;0.01 0.32&#b1;0.01 <0.0001 ߓ Cholestanol/chol ratio 1.75&#b1;0.04 1.45&#b1;0.03 <0.0001 Plant stanols ߓCampestanol, bc;g/dL 5.86&#b1;0.20 4.74&#b1;0.18 <0.0001 ߓ Campestanol/chol ratio 0.03&#b1;0.001 0.02&#b1;0.001 <0.0001 ߓSitostanol, bc;g/dL 7.80&#b1;0.28 6.39&#b1;0.25 <0.0001 ߓ Sitostanol/chol ratio, bc;g/mg 0.04&#b1;0.001 0.03&#b1;0.001 <0.0001 Markers of cholesterol synthesis ߓLanosterol, bc;g/dL 15.68&#b1;0.74 18.15&#b1;0.69 0.0009 ߓ Lanosterol/chol ratio 0.08&#b1;0.003 0.08&#b1;0.003 0.007 ߓ Lathosterol, mg/dL 0.20&#b1;0.01 0.24&#b1;0.01 <0.0001 ߓ Lathosterol/chol ratio 0.95&#b1;0.05 1.12&#b1;0.04 0.0006 ߓ Desmosterol, mg/dL 0.17&#b1;0.007 0.18&#b1;0.006 0.09 ߓ Desmosterol/chol ratio 0.78&#b1;0.03 0.82&#b1;0.03 0.15 Markers of bile acid synthesis ߓ7 b1; cholesterol 71.3&#b1;4.5 83.9&#b1;4.3 0.005 ߓ7 b1; cholesterol/chol ratio 0.33&#b1;0.02 0.38&#b1;0.02 0.02 *Chol indicates total cholesterol; and sitosterol/chol ratio, ratio of sitosterol to total cholesterol levels. Login to comment
96 suggest that G574R carriers are at higher risk for subclinical atheroscelerosis in light of their moderately higher plant sterol levels. Login to comment
97 Yet, our results offer an intriguing suggestion that G574R carriers may actually experience lower risk. Login to comment
110 Mean (and SD) values of intima-media wall thickness (IMT, in mm) between ABCG8 G574R heterozygotes and G574G (noncarriers) (A) and carotid IMT values in heterozygotes and noncarriers by age (B). Login to comment
119 In conclusion, our study presents the first data on a large cohort of carriers of the ABCG8 G574R variant. Login to comment

[hide] ABCG5/ABCG8 in cholesterol excretion and atheroscl... Clin Chim Acta. 2014 Jan 20;428:82-8. doi: 10.1016/j.cca.2013.11.010. Epub 2013 Nov 16. Yu XH, Qian K, Jiang N, Zheng XL, Cayabyab FS, Tang CK
ABCG5/ABCG8 in cholesterol excretion and atherosclerosis.
Clin Chim Acta. 2014 Jan 20;428:82-8. doi: 10.1016/j.cca.2013.11.010. Epub 2013 Nov 16., [PMID:24252657]

Abstract [show]
Cholesterol is essential for the growth and function of all mammalian cells, but abnormally increased blood cholesterol is a major risk factor for atherosclerotic cardiovascular disease. ATP-binding cassette (ABC) transporters G5 (ABCG5) and G8 (ABCG8) form an obligate heterodimer that limits intestinal absorption and facilitates biliary secretion of cholesterol and phytosterols. Consistent with their function, ABCG5 and ABCG8 are located on the apical membrane of enterocytes and hepatocytes. Liver X receptor is the major positive regulator of ABCG5 and ABCG8 expression. Mutations in either of the two genes cause sitosterolemia, a condition in which cholesterol and plant sterols accumulate in the circulation leading to premature cardiovascular disease. Overexpression of ABCG5 and ABCG8 in mice retards diet-induced atherosclerosis because of reduced circulating and hepatic cholesterol. In the current review, we summarize recent developments and propose a future framework that provides new perspectives on the regulation of cholesterol metabolism and treatment of atherosclerotic cardiovascular disease.

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828 Horenstein et al. have demonstrated that although the G574R variant of ABCG8 is associated with moderately elevated plant sterol levels, carriers of the G574R allele have modestly thinner carotid wall thickness than noncarriers [63]. Login to comment

[hide] Premature atherosclerosis is not systematic in phy... Atherosclerosis. 2014 May;234(1):162-8. doi: 10.1016/j.atherosclerosis.2014.02.030. Epub 2014 Mar 11. Hansel B, Carrie A, Brun-Druc N, Leclert G, Chantepie S, Coiffard AS, Kahn JF, Chapman MJ, Bruckert E
Premature atherosclerosis is not systematic in phytosterolemic patients: severe hypercholesterolemia as a confounding factor in five subjects.
Atherosclerosis. 2014 May;234(1):162-8. doi: 10.1016/j.atherosclerosis.2014.02.030. Epub 2014 Mar 11., [PMID:24657386]

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OBJECTIVE: Phytosterolemia is a rare autosomal recessive disorder characterized by dramatically elevated circulating levels of plant sterols (PS). Phytosterolemia is believed to be responsible for severe premature atherosclerosis. The clinical, biological and molecular genetic features of 5 patients with phytosterolemia and transient severe hypercholesterolemia challenge this hypothesis. METHODS: Our patients were referred for suspected homozygous familial hypercholesterolemia. Despite the phenotype, this diagnosis was invalidated and phytosterolemia was confirmed by the identification of mutations in the ABCG5/ABCG8 transporter complex. Plasma PS were analyzed with a mass spectrometric-gas chromatographic procedure. Vascular status was assessed with carotid ultrasonography and completed (for 4 of the 5 patients) with femoral ultrasonography; additional examinations of cardiovascular status included a stress test, determination of coronary calcium score, echocardiography, non-invasive assessment of endothelium-dependent dilatation and coronarography. RESULTS: The 5 patients displayed markedly elevated levels of both beta-sitosterol and campesterol (15-30 fold higher than normal values). However, none displayed significant signs of infraclinical premature atherosclerosis (respectively at the ages of 32, 27, 29, 11 and 11 years). All patients were characterized by very high levels of total (>450 mg/dl) and LDL-cholesterol (>350 mg/dl) at diagnosis which decreased markedly on dietary intervention alone. Treatment with cholestyramine or Ezetimibe +/- atorvastatin normalized cholesterol levels, although plasma PS concentrations remained elevated. CONCLUSION: The clinical and biological characteristics of our patients, considered together with reports of cases which equally lack CVD, support the contention that the premature atherosclerosis associated with phytosterolemia in some patients may be due at least in part to mechanisms independent of elevated circulating phytosterol levels.

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204 They compared mean plasma levels of plant sterols and cholesterol precursors and carotid intima media wall thickness in old order Amish participants with and without one copy of the ABCG8 G574R variant. Login to comment

[hide] A functional steroid-binding element in an ATP-bin... Mol Pharmacol. 2008 Jan;73(1):12-7. Velamakanni S, Janvilisri T, Shahi S, van Veen HW
A functional steroid-binding element in an ATP-binding cassette multidrug transporter.
Mol Pharmacol. 2008 Jan;73(1):12-7., [PMID:18094074]

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The human breast cancer resistance protein is an ATP-binding cassette (ABC) multidrug transporter that affects the bioavailability of chemotherapeutic drugs and can confer drug resistance on cancer cells. It is the second member of the ABCG subfamily, other members of which are associated with human steroid disorders such as hypercholesterolemia, sitosterolemia, and atherosclerosis. The molecular bases of protein-steroid interactions in ABC transporters are unknown. Here, we identify a steroid-binding element in the membrane domain of ABCG2 with a similarity to steroid hormone/nuclear receptors. The element facilitates steroid hormone binding and mediates modulation of ABCG2 activity. The identification of this element might provide an opportunity for the development of new therapeutic ligands for ABCG2.

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69 By analogy to the sitosterolemia-associated G574R substitution in ABCG8, Gly553 in the SxxLxxL motif was replaced by arginine (GR mutant). Login to comment