title: A splicing variant leads to complete loss of function of betaine-homocysteine methyltransferase (BHMT) gene in hepatocellular carcinoma.
abstract: The remethylation of homocyteine into methionine is catalyzed either by methionine synthase (MTR) or by betaine-homocysteine methyltransferase (BHMT), in the liver. Choline/betaine deficiency and impaired BHMT pathway have been associated with hepatocellular carcinogenesis, in animal models. The molecular mechanisms that impair the BHMT pathway are unknown. We aimed to investigate BHMT, BHMT2, and MTR expression in HepG2 cells and human hepatocarcinoma tissues. Transcripts were quantified by RT-qPCR and splicing was assessed by analysis of exon junctions and sequencing of variants. Protein expression was studied by Western Blot, immunohistochemistry and enzyme activity. Tumor tissue was compared with surrounding healthy tissue. RT-qPCR of HepG2 cells and of tumor samples showed a strong decrease of transcripts of BHMT and BHMT2, compared to normal. MTR transcript levels were not different. The decreased BHMT expression resulted from the transcription of a splicing variant that produced a frameshift in exon 4, with a premature termination codon in exon 5 and a loss of function of the gene. This splicing variant did not fit with any mechanism resulting from known splicing consensus sequences and was not detected in normal adult and fetal liver. Consistently, BHMT activity was abolished in HepG2 and protein expression was not detectable in HepG2 and in 5 of the 6 tumor samples, compared to normal tissues. In conclusion, a transcription variant of exon 4 produces a loss of function of BHMT in human hepatocarcinoma. Whether this abnormal transcription of BHMT is part or consequence of liver carcinogenesis should deserve further investigations.
authors.affiliation: INSERM U954 Nutrition, Genetics, and Environmental Risk Exposure, Faculté de Médecine, Nancy Université, 9 Avenue de Forêt de Haye, 54500 Vandoeuvre Nancy, France.
authors.forename: Hélène
authors.initials: H
authors.lastname: Pellanda
authors.forename: Farès
authors.initials: F
authors.lastname: Namour
authors.forename: Ma'atem
authors.initials: M
authors.lastname: Fofou-Caillierez
authors.forename: Aude
authors.initials: A
authors.lastname: Bressenot
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Alberto
authors.forename: Céline
authors.initials: C
authors.lastname: Chéry
authors.forename: Ahmet
authors.initials: A
authors.lastname: Ayav
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Bronowicki
authors.forename: Jean-Louis
authors.initials: JL
authors.lastname: Guéant
authors.forename: Thierry
authors.initials: T
authors.lastname: Forges
mesh_terms: D017398:Alternative Splicing / Q000502:physiology*; D001622:Betaine / Q000378:metabolism; D050940:Betaine-Homocysteine S-Methyltransferase / Q000235:genetics* / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000201:enzymology / Q000235:genetics*; D045744:Cell Line, Tumor; D003404:Creatinine / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D048289:Imidazolidines / Q000378:metabolism; D008113:Liver Neoplasms / Q000201:enzymology / Q000235:genetics*; D008715:Methionine / Q000378:metabolism
chemical_list: D048289:Imidazolidines; D001622:Betaine; C037182:homocyclocreatine; D008715:Methionine; D003404:Creatinine; D050940:Betaine-Homocysteine S-Methyltransferase