Functional analysis and classification of homozygous and hypomorphic ABCA4 variants associated with Stargardt macular degeneration

Curtis, Susan B.; Molday, Laurie L.; Garces, Fabian A.; Molday, Robert S.

doi:10.1002/humu.24100

Cited by 24 publications

(37 citation statements)

References 59 publications

(111 reference statements)

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Section: Resultssupporting

confidence: 92%

“…In the absence of ATR, but in the presence of phosphatidylethanolamine (PE), WT ABCA4 displays basal activity which largely reflects the energy-dependent conformational change related to the flipping of the secondary substrate PE across membranes [15]. Addition of 40 µM ATR in the presence of PE leads to the formation of the primary substrate N-Ret-PE and a two-fold increase in ATPase activity in agreement with previous reports [13,14,19,28,29] ( Figure The ATPase activity of the TMD variants can be divided into three distinct groups. Group 1 includes variants that have basal ATPase activity below 50% of WT levels and show little or no N-Ret-PE-induced stimulation in ATPase activity; Group 2 comprises variants with basal ATPase activity between 50% and 80% of WT levels and shows modest N-Ret-PE-stimulated ATPase activity; and Group 3 consists of variants that have both basal and N-Ret-PE-stimulated ATPase activity comparable to WT.…”

Section: Functional Analysis Of Tmd Disease-associated Variants: Atpasupporting

confidence: 90%

“…* Classification is based equally on the relative expression and substrate-activated ATPase. For more details, see reference [ 19 ].…”

Section: Figurementioning

confidence: 99%

“…A number of disease-associated variants with amino acid substitutions in the exocytoplasmic domains (ECDs), the nucleotide-binding domains (NBDs), and the C-terminal segment of ABCA4 have been previously characterized [ 13 , 14 , 15 , 16 , 17 , 18 , 19 ]. Many of these variants result in a significant reduction in cellular expression, mislocalization of ABCA4, and a loss of functional properties including substrate-stimulated ATPase activity and ATP-dependent substrate transport activity.…”

Section: Introductionmentioning

confidence: 99%

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Functional Characterization of ABCA4 Missense Variants Linked to Stargardt Macular Degeneration

Garces

Scortecci

Molday

2020

IJMS

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ABCA4 is an ATP-binding cassette (ABC) transporter expressed in photoreceptors, where it transports its substrate, N-retinylidene-phosphatidylethanolamine (N-Ret-PE), across outer segment membranes to facilitate the clearance of retinal from photoreceptors. Mutations in ABCA4 cause Stargardt macular degeneration (STGD1), an autosomal recessive disorder characterized by a loss of central vision and the accumulation of bisretinoid compounds. The purpose of this study was to determine the molecular properties of ABCA4 variants harboring disease-causing missense mutations in the transmembrane domains. Thirty-eight variants expressed in culture cells were analyzed for expression, ATPase activities, and substrate binding. On the basis of these properties, the variants were divided into three classes: Class 1 (severe variants) exhibited significantly reduced ABCA4 expression and basal ATPase activity that was not stimulated by its substrate N-Ret-PE; Class 2 (moderate variants) showed a partial reduction in expression and basal ATPase activity that was modestly stimulated by N-Ret-PE; and Class 3 (mild variants) displayed expression and functional properties comparable to normal ABCA4. The p.R653C variant displayed normal expression and basal ATPase activity, but lacked substrate binding and ATPase activation, suggesting that arginine 653 contributes to N-Ret-PE binding. Our classification provides a basis for better understanding genotype–phenotype correlations and evaluating therapeutic treatments for STGD1.

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Section: Resultssupporting

confidence: 92%

Section: Functional Analysis Of Tmd Disease-associated Variants: Atpasupporting

confidence: 90%

“…* Classification is based equally on the relative expression and substrate-activated ATPase. For more details, see reference [ 19 ].…”

Section: Figurementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Functional Characterization of ABCA4 Missense Variants Linked to Stargardt Macular Degeneration

Garces

Scortecci

Molday

2020

IJMS

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“…For example, what’s the role of the ECD for ABCA4 function? A large number of disease-associated mutations in ECD indicate a vital functional role of ECD 44–47 . It has been suggested that the hydrophobic tunnel in the ECD of ABCA1 might serve as a potentially temporary storage or delivery passage for lipid substrates 20,48 .…”

Section: Discussionmentioning

confidence: 99%

Structural basis of substrate recognition and translocation by human ABCA4

Xie

Zhang

et al. 2021

Preprint

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Human ATP-binding cassette (ABC) subfamily A (ABCA) transporters mediate the transport of various lipid compounds across the membrane. Mutations in human ABCA transporters have been described to cause severe hereditary disorders associated with impaired lipid transport. However, little is known about the mechanistic details of substrate recognition and translocation by ABCA transporters. Here, we present three cryo-EM structures of human ABCA4, a retinal-specific ABCA transporter, in distinct functional states at resolutions of 3.3 to 3.4 angstrom. In the nucleotide-free state, the two transmembrane domains (TMDs) exhibited a lateral-opening conformation, allowing the lateral entry of substrate from the lipid bilayer. N-retinylidene-phosphatidylethanolamine (NRPE), the physiological lipid substrate of ABCA4, is sandwiched between the two TMDs in the luminal leaflet and is further stabilized by an extended loop from extracellular domain 1. In the ATP-bound state, the two TMDs displayed an unprecedented closed conformation, which precludes the substrate binding. Our study provides a molecular basis to understand the mechanism of ABCA4-mediated NRPE recognition and translocation, and suggests a common lateral access and extrusion mechanism for ABCA-mediated lipid transport.

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Association Between Genotype and Phenotype Severity in ABCA4-Associated Retinopathy

et al. 2023

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ImportanceABCA4-associated retinopathy is a common inherited retinal disease, and its phenotype spans from late-onset macular dystrophy to extensive cone-rod degeneration. Over 2000 disease-causing variants in the ABCA4 gene have been identified.ObjectiveTo investigate genotype-phenotype correlations in ABCA4-associated retinopathy.Design, Setting, and ParticipantsThis cohort study took place at a single referral center for inherited retinal diseases in Italy. Data were prospectively acquired from January 2015 to June 2022. Patients diagnosed with an inherited retinal disease related to biallelic ABCA4 variants were included for analysis.ExposureGenotype, classified into 4 groups according to the presence of the (1) p.Gly1961Glu allele, (2) a hypomorphic allele, (3) at least 1 moderate variant (moderate genotypes), or (4) 2 biallelic severe variants (severe genotypes).Main Outcomes and MeasuresTotal decreased autofluorescence (TDAF) and definitely decreased autofluorescence (DDAF) areas, inner and outer retinal volumes, and the respective progression rate.ResultsA total of 71 patients (median [IQR] age, 34 [22.4-47.2] years; 40 [56%] female) were included in the study, and 54 (76%) were followed up for a median (IQR) of 3.5 (1.6-4.7) years. Compared with moderate genotypes, those with the p.Gly1961Glu allele had smaller TDAF lesions by 61% (95% CI, −78% to −33%; P &lt; .001) and DDAF lesions by 77% (95% CI, −93% to −18%; P = .02), along with slower growth rates for both TDAF (0.05 mm/y; 95% CI, 0.01-0.07; P &lt; .001) and DDAF (0.06 mm/y; 95% CI, 0-0.12; P = .004). Hypomorphic alleles were associated with a thicker inner (+0.19 mm3; 95% CI, +0.02 to +0.36; P = .03) and outer retinal volume (+0.16 mm3; 95% CI, +0.03 to +0.28; P = .01) compared with moderate genotypes as well as a slower TDAF growth rate (0.05 mm/y; 95% CI, 0.01-0.08; P = .007). Severe genotypes had a 7-fold larger TDAF area (95% CI, 3.4-14.7; P &lt; .001) and 11-fold larger DDAF area (95% CI, 2.9-42.1; P &lt; .001) compared with moderate genotypes, along with faster growth rates estimated at 0.16 mm/y for TDAF (95% CI, 0.12-0.20; P &lt; .001) and 0.17 mm/y for DDAF (95% CI, 0.12-0.23; P &lt; .001).Conclusions and RelevanceIn this study of ABCA4-associated retinopathy, a 4-tier classification of genotypes was found to capture substantial variation in disease phenotype severity. These findings could prove beneficial for the prognostication of patients and warrant consideration of genotype in the design of future clinical trials.

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Functional analysis and classification of homozygous and hypomorphic ABCA4 variants associated with Stargardt macular degeneration

Cited by 24 publications

References 59 publications

Functional Characterization of ABCA4 Missense Variants Linked to Stargardt Macular Degeneration

Functional Characterization of ABCA4 Missense Variants Linked to Stargardt Macular Degeneration

Structural basis of substrate recognition and translocation by human ABCA4

Association Between Genotype and Phenotype Severity in ABCA4-Associated Retinopathy

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