Functional rescue of mutant ABCA1 proteins by sodium 4-phenylbutyrate

Sorrenson, Brie; Suetani, Rachel J.; Williams, Michael J.A.; Bickley, Vivienne M.; George, Peter M.; Jones, Gregory T.; McCormick, Sally P.A.

doi:10.1194/jlr.m027193

Cited by 30 publications

(30 citation statements)

References 27 publications

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“…Since the retention of mutant protein in the ER is often a consequence of incorrect protein folding [37], we wanted to evaluate the effects of two chemical chaperones known to aid in protein folding, 4-PBA and TUDCA [24,25,38-43] , on trafficking of the Bmpr2ΔEx2 mutant product to the cell surface. Bmpr2 ΔEx2/+ ciPECs treated with increasing concentrations of 4-PBA show a dose-dependent increase expression of the 130kDa Bmpr2ΔEx2 mutant product in the streptavidin pull-down (Figure 4A/B).…”

Section: Resultsmentioning

confidence: 99%

“…Cell surface biotinylation and N-glycosidase sensitivity assays indicate that the Bmpr2ΔEx2 mutant product does not traffic correctly to the cell surface and is retained in the ER. Additionally, chemical chaperones 4-PBA and TUDCA, agents that are known to aid in protein folding [24,25,38-43], partially restore trafficking of the Bmpr2ΔEx2 mutant product to the cell surface. These findings suggest that the Bmpr2ΔEx2 is retained in the ER as a result of mis-folding of the mutant product.…”

Section: Discussionmentioning

confidence: 99%

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Abnormal Trafficking of Endogenously Expressed BMPR2 Mutant Allelic Products in Patients with Heritable Pulmonary Arterial Hypertension

et al. 2013

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More than 200 heterozygous mutations in the type 2 BMP receptor gene, BMPR2, have been identified in patients with Heritable Pulmonary Arterial Hypertension (HPAH). More severe clinical outcomes occur in patients with BMPR2 mutations by-passing nonsense-mediated mRNA decay (NMD negative mutations). These comprise 40% of HPAH mutations and are predicted to express BMPR2 mutant products. However expression of endogenous NMD negative BMPR2 mutant products and their effect on protein trafficking and signaling function have never been described. Here, we characterize the expression and trafficking of an HPAH-associated NMD negative BMPR2 mutation that results in an in-frame deletion of BMPR2 EXON2 (BMPR2ΔEx2) in HPAH patient-derived lymphocytes and in pulmonary endothelial cells (PECs) from mice carrying the same in-frame deletion of Exon 2 (Bmpr2 ΔEx2/+ mice). The endogenous BMPR2ΔEx2 mutant product does not reach the cell surface and is retained in the endoplasmic reticulum. Moreover, chemical chaperones 4-PBA and TUDCA partially restore cell surface expression of Bmpr2ΔEx2 in PECs, suggesting that the mutant product is mis-folded. We also show that PECs from Bmpr2 ΔEx2/+ mice have defects in the BMP-induced Smad1/5/8 and Id1 signaling axis, and that addition of chemical chaperones restores expression of the Smad1/5/8 target Id1. These data indicate that the endogenous NMD negative BMPRΔEx2 mutant product is expressed but has a folding defect resulting in ER retention. Partial correction of this folding defect and restoration of defective BMP signaling using chemical chaperones suggests that protein-folding agents could be used therapeutically in patients with these NMD negative BMPR2 mutations.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Abnormal Trafficking of Endogenously Expressed BMPR2 Mutant Allelic Products in Patients with Heritable Pulmonary Arterial Hypertension

et al. 2013

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“…With several members implicated in various severe human diseases, the A subclass of mammalian ABC transporters can serve as an exemplary protein family still lacking sufficient structural description despite significant advances in biochemical and clinical characterization [7, 42, 43]. The intricate membrane topology, multiple posttranslational modifications and unusual size (Figure 1B) dictate the predominant choice of mammalian expression systems for production of these proteins, as utilized in a number of studies (see [14, 17–23, 44, 45] for some examples). Alternatively, the presence of large regions apparently capable of forming soluble individual domains (Figure 1B) prompted other researchers to produce these as isolated proteins in E. coli , an attractive host that facilitates obtaining large amounts of a target protein at a low cost [24–30, 46].…”

Section: Discussionmentioning

confidence: 99%

Expression, purification and structural properties of ABC transporter ABCA4 and its individual domains

Tsybovsky

Palczewski

2014

Protein Expression and Purification

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ABCA4 is a member of the A subfamily of ATP-binding cassette transporters that consists of large integral membrane proteins implicated in inherited human diseases. ABCA4 assists in the clearance of N-retinylidene-phosphatidylethanolamine, a potentially toxic by-product of the visual cycle formed in photoreceptors during light perception. Structural and functional studies of this protein have been hindered by its large size, membrane association, and domain complexity. Although mammalian, insect and bacterial systems have been used for expression of ABCA4 and its individual domains, the structural relevance of resulting proteins to the native transporter has yet to be established. We produced soluble domains of ABCA4 in E. coli and S. cerevisiae and the full-length transporter in HEK293 cells. Electron microscopy and size exclusion chromatography were used to assess the conformational homogeneity and structure of these proteins. We found that isolated ABCA4 domains formed large, heterogeneous oligomers cross-linked with non-specific disulphide bonds. Incomplete folding of cytoplasmic domain 2 was proposed based on fluorescence spectroscopy results. In contrast, full-length human ABCA4 produced in mammalian cells was found structurally equivalent to the native protein obtained from bovine photoreceptors. These findings offer recombinantly expressed full-length ABCA4 as an appropriate object for future detailed structural and functional characterization.

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“…The ATPbinding cassette transporter A1 is involved in HDL cholesterol deficiency and mutated versions of this gene have been implicated in cardiovascular disease. Evidence suggests that 4-PBA is capable of restoring plasma membrane localization of this protein and subsequently enhancing cholesterol efflux function in vitro and ex vivo (Sorrenson et al, 2013).…”

Section: The Effects Of 4-pba On Protein Misfolding Diseases -Geneticmentioning

confidence: 99%

The therapeutic effects of 4-phenylbutyric acid in maintaining proteostasis

Kolb

Ayaub

Zhou

et al. 2015

The International Journal of Biochemistry & Cell Biology

211

180

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Functional rescue of mutant ABCA1 proteins by sodium 4-phenylbutyrate

Cited by 30 publications

References 27 publications

Abnormal Trafficking of Endogenously Expressed BMPR2 Mutant Allelic Products in Patients with Heritable Pulmonary Arterial Hypertension

Abnormal Trafficking of Endogenously Expressed BMPR2 Mutant Allelic Products in Patients with Heritable Pulmonary Arterial Hypertension

Expression, purification and structural properties of ABC transporter ABCA4 and its individual domains

The therapeutic effects of 4-phenylbutyric acid in maintaining proteostasis

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