title: Heterologous Expression in Yeast of Human Ornithine Carriers ORNT1 and ORNT2 and of ORNT1 Alleles Implicated in HHH Syndrome in Humans.
abstract: Hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is an autosomal recessive metabolic disorder usually presenting in the neonatal period with intermittent episodes of hyperammonemia, psychomotor delay, and progressive encephalopathy. Adult cases usually evolve into frank spastic paraparesis. The syndrome is caused by mutations in SLC25A15/ORNT1 encoding the mitochondrial ornithine transporter; a second ornithine transporter, ORNT2 of unknown function, is also present in most placental mammals. ORNT2 is believed to originate from an ancient retro-transposition event. In yeast Saccharomyces cerevisiae the major function of the transporter (encoded by Arg11) is to shuttle ornithine from the mitochondrial matrix to the cytosol. Its inactivation abolishes growth in the absence of arginine.In this work, we used functional complementation in S. cerevisiae to characterize the function of human ORNT2 and to test the pathogenicity of ORNT1 mutations found in HHH patients. Notably, we found that human ORNT1 but not ORNT2 complements the deletion of the yeast gene, despite their high level of homology. However, we identified some key residues in ORNT2, which may recover its functional competence when replaced with the corresponding residues of ORNT1, suggesting that roles of the two transporters are different. Moreover, we used this system to test a series of missense mutations of ORNT1 identified in patients with HHH syndrome. All mutations had a detrimental effect on the functionality of the human gene, without however clear genotype-phenotype correlations. Our data support yeast as a simple and effective model to validate missense mutations occurring in patients with HHH.
authors.affiliation: Clinical genetics Unit, Department of Woman and Child Health, University of Padova, Via Giustiniani 3, 35128, Padova, Italy.|IRP "Città della Speranza", Padova, Italy.
authors.forename: Mara
authors.initials: M
authors.lastname: Doimo
authors.affiliation: Department of Biomedical Sciences, University of Padova, Padova, Italy.
authors.forename: Raffaele
authors.initials: R
authors.lastname: Lopreiato
authors.affiliation: Clinical genetics Unit, Department of Woman and Child Health, University of Padova, Via Giustiniani 3, 35128, Padova, Italy.|IRP "Città della Speranza", Padova, Italy.
authors.forename: Valentina
authors.initials: V
authors.lastname: Basso
authors.affiliation: Clinical genetics Unit, Department of Woman and Child Health, University of Padova, Via Giustiniani 3, 35128, Padova, Italy.|IRP "Città della Speranza", Padova, Italy.
authors.forename: Raissa
authors.initials: R
authors.lastname: Bortolotto
authors.affiliation: Molecular Medicine Unit, IRCCS Stella Maris, Pisa, Italy.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Tessa
authors.affiliation: Molecular Medicine Unit, IRCCS Stella Maris, Pisa, Italy.
authors.forename: Filippo M
authors.initials: FM
authors.lastname: Santorelli
authors.affiliation: Clinical genetics Unit, Department of Woman and Child Health, University of Padova, Via Giustiniani 3, 35128, Padova, Italy.|IRP "Città della Speranza", Padova, Italy.
authors.forename: Eva
authors.initials: E
authors.lastname: Trevisson
authors.affiliation: Clinical genetics Unit, Department of Woman and Child Health, University of Padova, Via Giustiniani 3, 35128, Padova, Italy. leonardo.salviati@unipd.it.|IRP "Città della Speranza", Padova, Italy. leonardo.salviati@unipd.it.
authors.forename: Leonardo
authors.initials: L
authors.lastname: Salviati
keywords: HHH; ORNT1; ORNT2; Ornithine; Urea cycle disorder; Yeast model