title: Distinct roles of galactose-1P in galactose-mediated growth arrest of yeast deficient in galactose-1P uridylyltransferase (GALT) and UDP-galactose 4'-epimerase (GALE).
abstract: Galactose is metabolized in humans and other species by the three-enzyme Leloir pathway comprised of galactokinase (GALK), galactose 1-P uridylyltransferase (GALT), and UDP-galactose 4'-epimerase (GALE). Impairment of GALT or GALE in humans results in the potentially lethal disorder galactosemia, and loss of either enzyme in yeast results in galactose-dependent growth arrest of cultures despite the availability of an alternate carbon source. In contrast, loss of GALK in humans is not life-threatening, and in yeast has no impact on the growth of cultures challenged with galactose. Further, the growth of both GALT-null and GALE-null yeast challenged with galactose is rescued by loss of GALK, thereby implicating the GALK reaction product, gal-1P, for a role in the galactose-sensitivity of both strains. However, the nature of that relationship has remained unclear. Here we have developed and applied a doxycycline-repressible allele of galactokinase to define the quantitative relationship between galactokinase activity, gal-1P accumulation, and growth arrest of galactose-challenged GALT or GALE-deficient yeast. Our results demonstrate a clear threshold relationship between gal-1P accumulation and galactose-mediated growth arrest in both GALT-null and GALE-null yeast, however, the threshold for the two strains is distinct. Further, we tested the galactose-sensitivity of yeast double-null for GALT and GALE, and found that although loss of GALT barely changed accumulation of gal-1P, it significantly lowered the accumulation of UDP-gal, and also dramatically rescued growth of the GALE-null cells. Together, these data suggest that while gal-1P alone may account for the galactose-sensitivity of GALT-null cells, other factors, likely to include UDP-gal accumulation, must contribute to the galactose-sensitivity of GALE-null cells.
authors.affiliation: Department of Human Genetics, Emory University School of Medicine, Room 325.2 Whitehead Building, 615 Michael Street, Atlanta, GA, USA.
authors.forename: Jane Odhiambo
authors.initials: JO
authors.lastname: Mumma
authors.forename: Juliet S
authors.initials: JS
authors.lastname: Chhay
authors.forename: Kerry L
authors.initials: KL
authors.lastname: Ross
authors.forename: Jana S
authors.initials: JS
authors.lastname: Eaton
authors.forename: Karen A
authors.initials: KA
authors.lastname: Newell-Litwa
authors.forename: Judith L
authors.initials: JL
authors.lastname: Fridovich-Keil
mesh_terms: D004318:Doxycycline / Q000494:pharmacology; D005686:Galactokinase / Q000235:genetics / Q000378:metabolism; D005690:Galactose / Q000378:metabolism; D005695:Galactosephosphates / Q000378:metabolism*; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects; D005800:Genes, Fungal; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D014534:UDPglucose 4-Epimerase / Q000235:genetics / Q000378:metabolism*; D006599:UDPglucose-Hexose-1-Phosphate Uridylyltransferase / Q000235:genetics / Q000378:metabolism*
chemical_list: D005695:Galactosephosphates; C029973:galactose-1-phosphate; D005686:Galactokinase; D006599:UDPglucose-Hexose-1-Phosphate Uridylyltransferase; D014534:UDPglucose 4-Epimerase; D004318:Doxycycline; D005690:Galactose