Riboflavin, otherwise known as vitamin B2, is an essential dietary component and represents the precursor of flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), the redox enzymatic cofactors required for mitochondrial terminal metabolism and for the functionality of mitochondrial respiratory chain. Protein folding, ROS production and defense, as well as redox epigenetics also depend on cellular supply of FAD. FAD formation in different cells starts from riboflavin uptake, which occurs via specialized carrier-mediated processes which are supported by three specific members of the solute carrier family 52 (SLC52A), identified and named respectively RFVT1, RFVT2 and RFVT3. Once in the cell, riboflavin is converted to FMN by riboflavin kinase (RFK, EC2.7.1.26) and FMN is converted, in turn, to FAD by FAD synthase (FADS, EC 2.7.7.2), coded by human FLAD1 gene. Alternative splicing of the FLAD1 gene generates different hFADS isoforms, with different sub-cellular localization, most of them containing and a C-terminal 3-phosphoadenosine 5-phosphosulfate (PAPS) reductase domain, which per se performs the FAD synthase activity and a N-terminal molybdopterin binding (MPTb) domain, which could perform a FAD hydrolase activity. Alterations of FAD synthesis and RFVT1 impairment have been correlated with a rare inherited neuro-muscular disorders, in some cases treatable with high doses of Riboflavin, named Multiple Acyl-CoA Dehydrogenase Deficiency (MADD, MIM: 231680). Here we describe a novel case of MADD caused by a truncating variant in FLAD1 resulting in a premature STOP codon in exon 2. Studies, carried out on patient fibroblasts, revealed a dramatic reduction in FADS protein level with corresponding reduction in FAD synthesis rate and in mitochondrial flavoenzyme amounts. The residual FAD activity measured could result from the expression of a novel isoform named 6, lacking MPTb domain, relevant for patient survival [1]. Immunoblotting analysis of RFVTs revealed a reduction in RFTV2 protein amount in fibroblasts of this patient, which is in line with the reduction of the total flavin cellular content observed by HPLC. The secondary defect of riboflavin transport rate in MADD patient, laying the foundations for explaining riboflavin therapy, links MADD to Brown-Vialetto-Van-Laere Syndrome (BVVLS, MIM: 21153; 614707) [2]. It is a neurological disorder characterized by bilateral sensoneural deafness, respiratory insufficiencies and progressive ponto-bulbar palsy, linked to mutations in RFVT2 and RFVT3 genes. A significant reduction in the intracellular levels of FMN and FAD was found in BVVLS patient fibroblasts grown in low extracellular riboflavin conditions [2]. We also pointed our attention on dysregulation of RFVTs expression linked to human colorectal [3] and some other types of cancer, generating a profound alteration of flavin cofactor homeostasis. Changing the level of RFVTs expression as a possible mean to reprogram cellular flavoproteome will be discussed.

Alteration of flavin homeostasis in neuromuscular disorders and cancer

Maria Tolomeo;Piero Leone;Maria Barile
2019

Abstract

Riboflavin, otherwise known as vitamin B2, is an essential dietary component and represents the precursor of flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), the redox enzymatic cofactors required for mitochondrial terminal metabolism and for the functionality of mitochondrial respiratory chain. Protein folding, ROS production and defense, as well as redox epigenetics also depend on cellular supply of FAD. FAD formation in different cells starts from riboflavin uptake, which occurs via specialized carrier-mediated processes which are supported by three specific members of the solute carrier family 52 (SLC52A), identified and named respectively RFVT1, RFVT2 and RFVT3. Once in the cell, riboflavin is converted to FMN by riboflavin kinase (RFK, EC2.7.1.26) and FMN is converted, in turn, to FAD by FAD synthase (FADS, EC 2.7.7.2), coded by human FLAD1 gene. Alternative splicing of the FLAD1 gene generates different hFADS isoforms, with different sub-cellular localization, most of them containing and a C-terminal 3-phosphoadenosine 5-phosphosulfate (PAPS) reductase domain, which per se performs the FAD synthase activity and a N-terminal molybdopterin binding (MPTb) domain, which could perform a FAD hydrolase activity. Alterations of FAD synthesis and RFVT1 impairment have been correlated with a rare inherited neuro-muscular disorders, in some cases treatable with high doses of Riboflavin, named Multiple Acyl-CoA Dehydrogenase Deficiency (MADD, MIM: 231680). Here we describe a novel case of MADD caused by a truncating variant in FLAD1 resulting in a premature STOP codon in exon 2. Studies, carried out on patient fibroblasts, revealed a dramatic reduction in FADS protein level with corresponding reduction in FAD synthesis rate and in mitochondrial flavoenzyme amounts. The residual FAD activity measured could result from the expression of a novel isoform named 6, lacking MPTb domain, relevant for patient survival [1]. Immunoblotting analysis of RFVTs revealed a reduction in RFTV2 protein amount in fibroblasts of this patient, which is in line with the reduction of the total flavin cellular content observed by HPLC. The secondary defect of riboflavin transport rate in MADD patient, laying the foundations for explaining riboflavin therapy, links MADD to Brown-Vialetto-Van-Laere Syndrome (BVVLS, MIM: 21153; 614707) [2]. It is a neurological disorder characterized by bilateral sensoneural deafness, respiratory insufficiencies and progressive ponto-bulbar palsy, linked to mutations in RFVT2 and RFVT3 genes. A significant reduction in the intracellular levels of FMN and FAD was found in BVVLS patient fibroblasts grown in low extracellular riboflavin conditions [2]. We also pointed our attention on dysregulation of RFVTs expression linked to human colorectal [3] and some other types of cancer, generating a profound alteration of flavin cofactor homeostasis. Changing the level of RFVTs expression as a possible mean to reprogram cellular flavoproteome will be discussed.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11586/347820
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