BACKGROUND: Remethylation defects are rare inherited disorders in which impaired remethylation of homocysteine to methionine leads to accumulation of homocysteine and perturbation of numerous methylation reactions. OBJECTIVE: To summarise clinical and biochemical characteristics of these severe disorders and to provide guidelines on diagnosis and management. DATA SOURCES: Review, evaluation and discussion of the medical literature (Medline, Cochrane databases) by a panel of experts on these rare diseases following the GRADE approach. KEY RECOMMENDATIONS: We strongly recommend measuring plasma total homocysteine in any patient presenting with the combination of neurological and/or visual and/or haematological symptoms, subacute spinal cord degeneration, atypical haemolytic uraemic syndrome or unexplained vascular thrombosis. We strongly recommend to initiate treatment with parenteral hydroxocobalamin without delay in any suspected remethylation disorder; it significantly improves survival and incidence of severe complications. We strongly recommend betaine treatment in individuals with MTHFR deficiency; it improves the outcome and prevents disease when given early.

• MeSH
• Homocysteine genetics   MeSH
• Humans MeSH
• Methionine genetics   MeSH
• Methylenetetrahydrofolate Reductase (NADPH2) deficiency   MeSH
• Methylation drug effects   MeSH
• Proto-Oncogene Proteins c-cbl deficiency   MeSH
• Vitamin B 12 pharmacology   therapeutic use   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Homocysteine MeSH
• Methionine MeSH
• Methylenetetrahydrofolate Reductase (NADPH2) MeSH
• Proto-Oncogene Proteins c-cbl MeSH
• Vitamin B 12 MeSH

Newborn screening (NBS) is justified if early intervention is effective in a disorder generally not detected early in life on a clinical basis, and if sensitive and specific biochemical markers exist. Experience with NBS for homocystinurias and methylation disorders is limited. However, there is robust evidence for the success of early treatment with diet, betaine and/or pyridoxine for CBS deficiency and good evidence for the success of early betaine treatment in severe MTHFR deficiency. These conditions can be screened in dried blood spots by determining methionine (Met), methionine-to-phenylanine (Met/Phe) ratio, and total homocysteine (tHcy) as a second tier marker. Therefore, we recommend NBS for cystathionine beta-synthase and severe MTHFR deficiency. Weaker evidence is available for the disorders of intracellular cobalamin metabolism. Early treatment is clearly of advantage for patients with the late-onset cblC defect. In the early-onset type, survival and non-neurological symptoms improve but the effect on neurocognitive development is uncertain. The cblC defect can be screened by measuring propionylcarnitine, propionylcarnitine-to-acetylcarnitine ratio combined with the second tier markers methylmalonic acid and tHcy. For the cblE and cblG defects, evidence for the benefit of early treatment is weaker; and data on performance of Met, Met/Phe and tHcy even more limited. Individuals homozygous or compound heterozygous for MAT1A mutations may benefit from detection by NBS using Met, which on the other hand also detects asymptomatic heterozygotes. Clinical and laboratory data is insufficient to develop any recommendation on NBS for the cblD, cblF, cblJ defects, glycineN-methyltransferase-, S-adenosylhomocysteinehydrolase- and adenosine kinase deficiency.

• MeSH
• Acetylcarnitine blood   MeSH
• Betaine therapeutic use   MeSH
• Homocystinuria diagnosis   MeSH
• Carnitine analogs & derivatives   blood   MeSH
• Methylmalonic Acid blood   MeSH
• Humans MeSH
• Methionine blood   MeSH
• Methylenetetrahydrofolate Reductase (NADPH2) deficiency   drug effects   MeSH
• Methylation MeSH
• Infant, Newborn MeSH
• Neonatal Screening * MeSH
• Practice Guidelines as Topic MeSH
• Check Tag
• Humans MeSH
• Infant, Newborn MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Systematic Review MeSH
• Names of Substances
• Acetylcarnitine MeSH
• Betaine MeSH
• Carnitine MeSH
• Methylmalonic Acid MeSH
• Methionine MeSH
• Methylenetetrahydrofolate Reductase (NADPH2) MeSH
• propionylcarnitine MeSH Browser

The prevalent c.903+469T>C mutation in MTRR causes the cblE type of homocystinuria by strengthening an SRSF1 binding site in an ESE leading to activation of a pseudoexon. We hypothesized that other splicing regulatory elements (SREs) are also critical for MTRR pseudoexon inclusion. We demonstrate that the MTRR pseudoexon is on the verge of being recognized and is therefore vulnerable to several point mutations that disrupt a fine-tuned balance between the different SREs. Normally, pseudoexon inclusion is suppressed by a hnRNP A1 binding exonic splicing silencer (ESS). When the c.903+469T>C mutation is present two ESEs abrogate the activity of the ESS and promote pseudoexon inclusion. Blocking the 3'splice site or the ESEs by SSOs is effective in restoring normal splicing of minigenes and endogenous MTRR transcripts in patient cells. By employing an SSO complementary to both ESEs, we were able to rescue MTRR enzymatic activity in patient cells to approximately 50% of that in controls. We show that several point mutations, individually, can activate a pseudoexon, illustrating that this mechanism can occur more frequently than previously expected. Moreover, we demonstrate that SSO blocking of critical ESEs is a promising strategy to treat the increasing number of activated pseudoexons.

• MeSH
• Cell Line MeSH
• Exons * MeSH
• Ferredoxin-NADP Reductase genetics   metabolism   MeSH
• HEK293 Cells MeSH
• Homocystinuria enzymology   genetics   MeSH
• Cells, Cultured MeSH
• Humans MeSH
• Anemia, Megaloblastic enzymology   genetics   MeSH
• RNA Splice Sites MeSH
• Mutation * MeSH
• Oligonucleotides * MeSH
• Regulatory Sequences, Ribonucleic Acid * MeSH
• RNA Splicing * MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Ferredoxin-NADP Reductase MeSH
• methionine synthase reductase MeSH Browser
• RNA Splice Sites MeSH
• Oligonucleotides * MeSH
• Regulatory Sequences, Ribonucleic Acid * MeSH

Deep intronic mutations are often ignored as possible causes of human diseases. A deep intronic mutation in the MTRR gene, c.903+469T>C, is the most frequent mutation causing the cblE type of homocystinuria. It is well known to be associated with pre-mRNA mis-splicing, resulting in pseudoexon inclusion; however, the pathological mechanism remains unknown. We used minigenes to demonstrate that this mutation is the direct cause of MTRR pseudoexon inclusion, and that the pseudoexon is normally not recognized due to a suboptimal 5' splice site. Within the pseudoexon we identified an exonic splicing enhancer (ESE), which is activated by the mutation. Cotransfection and siRNA experiments showed that pseudoexon inclusion depends on the cellular amounts of SF2/ASF and in vitro RNA-binding assays showed dramatically increased SF2/ASF binding to the mutant MTRR ESE. The mutant MTRR ESE sequence is identical to an ESE of the alternatively spliced MST1R proto-oncogene, which suggests that this ESE could be frequently involved in splicing regulation. Our study conclusively demonstrates that an intronic single nucleotide change is sufficient to cause pseudoexon activation via creation of a functional ESE, which binds a specific splicing factor. We suggest that this mechanism may cause genetic disease much more frequently than previously reported.

• MeSH
• Chlorocebus aethiops MeSH
• COS Cells MeSH
• Exons genetics   MeSH
• Ferredoxin-NADP Reductase genetics   MeSH
• Homocystinuria classification   enzymology   genetics   MeSH
• Introns genetics   MeSH
• Nuclear Proteins metabolism   MeSH
• RNA, Messenger genetics   metabolism   MeSH
• RNA Splice Sites genetics   MeSH
• Molecular Sequence Data MeSH
• Mutation genetics   MeSH
• Mutant Proteins genetics   MeSH
• RNA-Binding Proteins metabolism   MeSH
• Proto-Oncogene Mas MeSH
• Base Sequence MeSH
• Serine-Arginine Splicing Factors MeSH
• RNA Splicing genetics   MeSH
• Protein Binding MeSH
• Vitamin B 12 metabolism   MeSH
• Computational Biology MeSH
• Enhancer Elements, Genetic genetics   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Ferredoxin-NADP Reductase MeSH
• Nuclear Proteins MeSH
• MAS1 protein, human MeSH Browser
• RNA, Messenger MeSH
• methionine synthase reductase MeSH Browser
• RNA Splice Sites MeSH
• Mutant Proteins MeSH
• RNA-Binding Proteins MeSH
• Proto-Oncogene Mas MeSH
• Serine-Arginine Splicing Factors MeSH
• Vitamin B 12 MeSH