title: Humanizing mouse folate metabolism: conversion of the dual-promoter mouse folylpolyglutamate synthetase gene to the human single-promoter structure.
abstract: The mouse is extensively used to model human folate metabolism and therapeutic outcomes with antifolates. However, the folylpoly-γ-glutamate synthetase (fpgs) gene, whose product determines folate/antifolate intracellular retention and antifolate antitumor activity, displays a pronounced species difference. The human gene uses only a single promoter, whereas the mouse uses two: P2, akin to the human promoter, at low levels in most tissues; and P1, an upstream promoter used extensively in liver and kidney. We deleted the mouse P1 promoter through homologous recombination to study the dual-promoter mouse system and to create a mouse with a humanized fpgs gene structure. Despite the loss of the predominant fpgs mRNA species in liver and kidney (representing 95 and 75% of fpgs transcripts in these tissues, respectively), P1-knockout mice developed and reproduced normally. The survival of these mice was explained by increased P2 transcription due to relief of transcriptional interference, by a 3-fold more efficient translation of P2-derived than P1-derived transcripts, and by 2-fold higher stability of P2-derived FPGS. In combination, all 3 effects reinstated FPGS function, even in liver. By eliminating mouse P1, we created a mouse model that mimicked the human housekeeping pattern of fpgs gene expression.
authors.affiliation: 2Massey Cancer Center, Virginia Commonwealth University, 401 College St., Richmond, VA 23298-0035, USA. rmoran@vcu.edu.
authors.forename: Chen
authors.initials: C
authors.lastname: Yang
authors.forename: Lin-Ying
authors.initials: LY
authors.lastname: Xie
authors.forename: Jolene J
authors.initials: JJ
authors.lastname: Windle
authors.forename: Shirley M
authors.initials: SM
authors.lastname: Taylor
authors.forename: Richard G
authors.initials: RG
authors.lastname: Moran
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D001483:Base Sequence; D053595:Embryonic Stem Cells / Q000166:cytology; D005091:Exons; D005492:Folic Acid / Q000378:metabolism*; D005493:Folic Acid Antagonists / Q000494:pharmacology; D017353:Gene Deletion; D020869:Gene Expression Profiling; D005822:Genetic Vectors; D006720:Homozygote; D006801:Humans; D008099:Liver / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D010453:Peptide Synthases / Q000235:genetics*; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D021381:Protein Transport; D011995:Recombination, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
chemical_list: D000970:Antineoplastic Agents; D005493:Folic Acid Antagonists; D005492:Folic Acid; D010453:Peptide Synthases; C023103:folylpolyglutamate synthetase
keywords: knockout mice; protein translocation and degradation; transcription; transcriptional interference; translation