The clinically relevant MTARC1 p.Ala165Thr variant impacts neither the fold nor active site architecture of the human mARC1 protein

Struwe, Michel A.; Clement, Bernd; Scheidig, Axel J.

doi:10.1101/2022.03.26.485076

Cited by 4 publications

(6 citation statements)

References 18 publications

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“…The structure has the PDB accession code 7P41, and experimental details are also publicly available. [ 5 ] Briefly, while the electron density of the mutated residue is well‐defined, no other alterations of the structure are observed: The alpha helix predicted to be lost in the variant is fully intact, as is the molybdopterin cofactor and the molybdenum ion's coordination environment (Figure 1 ).…”

Section: Figurementioning

confidence: 99%

Letter to the editor: The clinically relevant MTARC1 p.Ala165Thr variant impacts neither the fold nor active site architecture of the human mARC1 protein

2022

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Section: Figurementioning

confidence: 99%

Letter to the editor: The clinically relevant MTARC1 p.Ala165Thr variant impacts neither the fold nor active site architecture of the human mARC1 protein

2022

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“…Previous studies indicated that the protective loss-of-function MTARC1 variants identified by genetics do not affect enzymatic activity or protein folding. 18 19 We reasoned that the protective effect of these variants must be mediated by some other mechanism such as protein stability or subcellular localization. Consequently, we studied the localization and protein accumulation of 3 published human missense and nonsense variants associated with protection from MASH (A allele of rs2642438, p.M187K, and p.R200*), 11 as well as one synthetic mutation known to abrogate the enzymatic function of the protein, p.C273A 20 using in vitro overexpression experiments.…”

Section: Resultsmentioning

confidence: 99%

mARC1 in MASLD: Modulation of lipid accumulation in human hepatocytes and adipocytes

Jones,

Bajrami,

Campbell

et al. 2024

Hepatology Communications

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Background: Mutations in the gene MTARC1 (mitochondrial amidoxime–reducing component 1) protect carriers from metabolic dysfunction–associated steatohepatitis (MASH) and cirrhosis. MTARC1 encodes the mARC1 enzyme, which is localized to the mitochondria and has no known MASH-relevant molecular function. Our studies aimed to expand on the published human genetic mARC1 data and to observe the molecular effects of mARC1 modulation in preclinical MASH models. Methods and Results: We identified a novel human structural variant deletion in MTARC1, which is associated with various biomarkers of liver health, including alanine aminotransferase levels. Phenome-wide Mendelian Randomization analyses additionally identified novel putatively causal associations between MTARC1 expression, and esophageal varices and cardiorespiratory traits. We observed that protective MTARC1 variants decreased protein accumulation in in vitro overexpression systems and used genetic tools to study mARC1 depletion in relevant human and mouse systems. Hepatocyte mARC1 knockdown in murine MASH models reduced body weight, liver steatosis, oxidative stress, cell death, and fibrogenesis markers. mARC1 siRNA treatment and overexpression modulated lipid accumulation and cell death consistently in primary human hepatocytes, hepatocyte cell lines, and primary human adipocytes. mARC1 depletion affected the accumulation of distinct lipid species and the expression of inflammatory and mitochondrial pathway genes/proteins in both in vitro and in vivo models. Conclusions: Depleting hepatocyte mARC1 improved metabolic dysfunction–associated steatotic liver disease–related outcomes. Given the functional role of mARC1 in human adipocyte lipid accumulation, systemic targeting of mARC1 should be considered when designing mARC1 therapies. Our data point to plasma lipid biomarkers predictive of mARC1 abundance, such as Ceramide 22:1. We propose future areas of study to describe the precise molecular function of mARC1, including lipid trafficking and subcellular location within or around the mitochondria and endoplasmic reticulum.

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“…Evidence from mARC1 crystal structures and published data showing similar molybdenum content among different recombinant mARC1 variants both demonstrate that residue 165 is not involved in Moco-binding (30,39). A recent crystal structure study of mARC1 A165T showed that there were alternate conformations of Thr165, even though no difference in protein folding and thermal stability was found between A165T and WT (40).…”

Section: Discussionmentioning

confidence: 99%

Biochemical and functional characterization of the p.A165T missense variant of mitochondrial amidoxime-reducing component 1 in HepG2 cells

Hou,

Watson,

Cecconie

et al. 2023

Preprint

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Recent genome-wide association studies have identified a missense variant p.A165T in mitochondrial amidoxime-reducing component 1 (mARC1) that is strongly associated with protection from all-cause cirrhosis and improved prognosis in nonalcoholic steatohepatitis (NASH). The precise mechanism of this protective effect is unknown. Substitution of alanine 165 with threonine is predicted to affect mARC1 protein stability and to have deleterious effects on its function. To investigate the mechanism, we have generated a knock-in mutant mARC1 A165T in human hepatoma HepG2 cells, enabling characterization of protein subcellular distribution, stability, and biochemical functions of the mARC1 mutant protein expressed from its endogenous locus. Compared to wild-type (WT) mARC1, we found that the A165T mutant exhibits significant mislocalization outside of its traditional location anchored in the mitochondrial outer membrane and reduces protein stability, resulting in lower basal levels. We evaluated the involvement of the ubiquitin proteasome system in mARC1 A165T degradation and observed increased ubiquitination and faster degradation of the A165T variant. In addition, we have shown that HepG2 cells carrying the MTARC1 p.A165T variant exhibit lower N-reductive activity on exogenously-added amidoxime substrates in vitro. The data from these biochemical and functional assays suggest a mechanism by which the MTARC1 p.A165T variant abrogates enzyme function which may contribute to its protective effect in liver disease.

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The clinically relevant MTARC1 p.Ala165Thr variant impacts neither the fold nor active site architecture of the human mARC1 protein

Cited by 4 publications

References 18 publications

Letter to the editor: The clinically relevant MTARC1 p.Ala165Thr variant impacts neither the fold nor active site architecture of the human mARC1 protein

Letter to the editor: The clinically relevant MTARC1 p.Ala165Thr variant impacts neither the fold nor active site architecture of the human mARC1 protein

mARC1 in MASLD: Modulation of lipid accumulation in human hepatocytes and adipocytes

Biochemical and functional characterization of the p.A165T missense variant of mitochondrial amidoxime-reducing component 1 in HepG2 cells

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