Corneal Dystrophy Mutations Drive Pathogenesis by Targeting TGFBIp Stability and Solubility in a Latent Amyloid-forming Domain

Stenvang, Marcel; Schafer, Nicholas P.; Malmos, Kirsten Gade; Pérez, Adriana-Michelle Wolf; Niembro, Olatz; Sormanni, Pietro; Basaiawmoit, Rajiv Vaid; Christiansen, Gunna; Andreasen, Maria; Otzen, Daniel E.

doi:10.1016/j.jmb.2018.03.001

Cited by 18 publications

(23 citation statements)

References 40 publications

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“…To validate TGFBIp or some of its degradation products as HtrA1 substrates, the in vitro proteolysis of wild-type and mutant TGFBIp and FAS1-4 proteins was analyzed (Poulsen et al, 2019). HtrA1 showed the highest preferences for mutant TGFBIp associated with the LCD phenotype, especially when using the FAS1-4 domain as a substrate (consistent with the increased protease sensitivity of these mutants (Stenvang et al, 2018)). Furthermore, degraded FAS1-4 formed amyloid fibrils consisting mainly of peptides overlapping with the Y571-R588 region highly represented in in vivo LCD deposits.…”

Section: Htra1 Proteolysis In Lcdmentioning

confidence: 74%

“…Of these, both LCD mutants and the variant GCD2 mutant were destabilizing; these mutants also exposed more hydrophobic surface area (i.e., were "sticky") and readily succumbed to trypsin degradation. Remarkably, despite a similar lack of stability, the LCD mutants formed amyloid significantly better than their GCD2 counterpart, and the aggregates formed by the variant GCD2 mutant (like the GCD1 mutant R555W) were non-amyloid (Stenvang et al, 2018).…”

Section: A Comprehensive Mutagenic and Bioinformatics Study Highlights Fundamental Differences Between Lcd And Gcd Mutantsmentioning

confidence: 94%

“…Based on this overall confirmation of the destabilizing properties of LCD mutants, we carried out a comprehensive computational analysis of the known and predicted properties of all known FAS1-4 mutations associated with TGFBI-linked corneal dystrophies and reached some striking conclusions (Stenvang et al, 2018).…”

Section: A Comprehensive Mutagenic and Bioinformatics Study Highlights Fundamental Differences Between Lcd And Gcd Mutantsmentioning

confidence: 97%

“…Building on the connection between FAS1-4 stability and phenotype (Runager et al, 2011), we systematically probed the link between the clinical manifestations of TGFBI-linked corneal dystrophy mutations and their underlying molecular impact on protein folding and stability. Our study encompassed 30 FAS1-4 mutants, spanning both LCD as well as GCD1, variant GCD2, and TBCD (Stenvang et al, 2018). Recombinantly expressed mutants (which included a tryptophan residue introduced to aid spectroscopic measurements) were analyzed in terms of their tendency to oligomerize in solution, stability against urea and thermal denaturation, tendency to expose hydrophobic surface area, sensitivity to protease digestion, protein folding/unfolding dynamics, and tendency to form amyloid in the presence of the glycosaminoglycan (GAG) heparin.…”

Section: A Comprehensive Mutagenic and Bioinformatics Study Highlights Fundamental Differences Between Lcd And Gcd Mutantsmentioning

confidence: 99%

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Biochemical mechanisms of aggregation in TGFBI-linked corneal dystrophies

Nielsen

Poulsen

Lukassen

et al. 2020

Progress in Retinal and Eye Research

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Transforming growth factor-β-induced protein (TGFBIp), an extracellular matrix protein, is the second most abundant protein in the corneal stroma. In this review, we summarize the current knowledge concerning the expression, molecular structure, binding partners, and functions of human TGFBIp. To date, 74 mutations in the transforming growth factor-β-induced gene (TGFBI) are associated with amyloid and amorphous protein deposition in TGFBI-linked corneal dystrophies. We discuss the current understanding of the biochemical mechanisms of TGFBI-linked corneal dystrophies and propose that mutations leading to granular corneal dystrophy (GCD) decrease the solubility of TGFBIp and affect the interactions between TGFBIp and components of the corneal stroma, whereas mutations associated with lattice corneal dystrophy (LCD) lead to a destabilization of the protein that disrupts proteolytic turnover, especially by the serine protease HtrA1. Future research should focus on TGFBIp function in the cornea, confirmation of the biochemical mechanisms in vivo, and the development of disease models. Future therapies for TGFBI-linked corneal dystrophies might include topical agents that regulate protein aggregation or gene therapy that targets the mutant allele by CRISPR/Cas9 technology.

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Section: Htra1 Proteolysis In Lcdmentioning

confidence: 74%

Section: A Comprehensive Mutagenic and Bioinformatics Study Highlights Fundamental Differences Between Lcd And Gcd Mutantsmentioning

confidence: 94%

Section: A Comprehensive Mutagenic and Bioinformatics Study Highlights Fundamental Differences Between Lcd And Gcd Mutantsmentioning

confidence: 97%

Section: A Comprehensive Mutagenic and Bioinformatics Study Highlights Fundamental Differences Between Lcd And Gcd Mutantsmentioning

confidence: 99%

See 2 more Smart Citations

Biochemical mechanisms of aggregation in TGFBI-linked corneal dystrophies

Nielsen

Poulsen

Lukassen

et al. 2020

Progress in Retinal and Eye Research

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“…The aggregation and deposition of TGFBIp display different clinical phenotypes; the deposits range from amyloidogenic structures to amorphous granular deposits, or a combination of both 17 . Mutations in TGFBIp are not only known to alter the turnover rate but also alter the thermodynamic stability of the protein with several of the mutations leading to destabilized protein which is more likely to unfold [18][19][20] . The mutant protein also possesses different proteolytic processing and clearance mechanism in the eye compared to the wild type protein (WT).…”

mentioning

confidence: 99%

Effect of osmolytes on in-vitro aggregation properties of peptides derived from TGFBIp

Venkatraman

Murugan

Lin

et al. 2020

Sci Rep

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Protein aggregation has been one of the leading triggers of various disease conditions, such asAlzheimer's, Parkinson's and other amyloidosis. TGFBI-associated corneal dystrophies are protein aggregation disorders in which the mutant TGFBIp aggregates and accumulates in the cornea, leading to a reduction in visual acuity and blindness in severe cases. Currently, the only therapy available is invasive and there is a known recurrence after surgery. In this study, we tested the inhibitory and amyloid dissociation properties of four osmolytes in an in-vitro TGFBI peptide aggregation model. The 23-amino acid long peptide (TGFBIp 611-633 with the mutation c.623 G>R) from the 4th FAS-1 domain of TGFBIp that rapidly forms amyloid fibrils was used in the study. Several biophysical methods like Thioflavin T (ThT) fluorescence, Circular Dichroism (CD), fluorescence microscopy and Transmission electron microscopy (TEM) were used to study the inhibitory and amyloid disaggregation properties of the four osmolytes (Betaine, Raffinose, Sarcosine, and Taurine). The osmolytes were effective in both inhibiting and disaggregating the amyloid fibrils derived from TGFBIp 611-633 c.623 G>R peptide. The osmolytes did not have an adverse toxic effect on cultured human corneal fibroblast cells and could potentially be a useful therapeutic strategy for patients with TGFBIp corneal dystrophies. Cytotoxicity of osmolytes on cultured human corneal fibroblast (HCF). Real-time live-cell imagingby IncuCyte. For real-time observation of the effect of the osmolytes on cultured human corneal fibroblast, 3000 cells/well were seeded in 96-well plates and placed in an incubator at 37 °C for 24 h to proliferate. Cells were then incubated with varying concentrations of the osmolytes (0.1 mM, 1 mM, 10 mM, 100 mM and 1000 mM) in triplicates and images were captured with IncuCyte ZOOM System (Essen BioScience Inc., Research Instruments, Singapore). Frames were then captured at 4-h intervals from 4 separate regions/well using a 10× objective for 20 h to observe cell toxicity. Scientific RepoRtS |(2020) 10:4011 | https://doi.

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Protein Analysis of the TGFBI^R124H Mouse Model Gives Insight into Phenotype Development of Granular Corneal Dystrophy

Lukassen

Poulsen

Donaghy

et al. 2020

Proteomics Clinical Apps

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Purpose: Mutations in the transforming growth factor-induced protein (TGFBIp) are associated with TGFBI-linked corneal dystrophies, which manifests as protein deposits in the cornea. A total of 70 different disease-causing mutations have been reported so far including the common R124H substitution, which is associated with granular corneal dystrophy type 2 (GCD2). The disease mechanism of GCD2 is not known and the current treatments only offer temporary relief due to the reoccurrence of deposits. Experimental Design: The corneal protein profiles of the three genotypes (wild-type (WT), heterozygotes, and homozygotes) of a GCD2 mouse model are compared using label-free quantitative LC-MS/MS. Results: The mice do not display corneal protein deposits and the global protein expression between the three genotypes is highly similar. However, the expression of mutated TGFBIp is 41% of that of the WT protein. Conclusions and Clinical Relevance: It is proposed that the lowered expression level of mutant TGFBIp protein relative to WT protein is the direct cause of the missing development of corneal deposits in the mouse. The overall protein profiles of the corneas are not impacted by the reduced amount of TGFBIp. Altogether, this supports a partial reduction in mutated TGFBIp as a potential treatment strategy for GCD2.

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Corneal Dystrophy Mutations Drive Pathogenesis by Targeting TGFBIp Stability and Solubility in a Latent Amyloid-forming Domain

Cited by 18 publications

References 40 publications

Biochemical mechanisms of aggregation in TGFBI-linked corneal dystrophies

Biochemical mechanisms of aggregation in TGFBI-linked corneal dystrophies

Effect of osmolytes on in-vitro aggregation properties of peptides derived from TGFBIp

Protein Analysis of the TGFBI^R124H Mouse Model Gives Insight into Phenotype Development of Granular Corneal Dystrophy

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Corneal Dystrophy Mutations Drive Pathogenesis by Targeting TGFBIp Stability and Solubility in a Latent Amyloid-forming Domain

Cited by 18 publications

References 40 publications

Biochemical mechanisms of aggregation in TGFBI-linked corneal dystrophies

Biochemical mechanisms of aggregation in TGFBI-linked corneal dystrophies

Effect of osmolytes on in-vitro aggregation properties of peptides derived from TGFBIp

Protein Analysis of the TGFBIR124H Mouse Model Gives Insight into Phenotype Development of Granular Corneal Dystrophy

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Protein Analysis of the TGFBI^R124H Mouse Model Gives Insight into Phenotype Development of Granular Corneal Dystrophy