Author: Cleary, M.D.; Meiering, C.D.; Jan, E.; Guymon, R.; Boothroyd, J.C.
Description: Standard microarrays measure mRNA abundance, not mRNA synthesis, and therefore cannot identify the mechanisms that regulate gene expression. We have developed a method to overcome this limitation by using the salvage enzyme uracil phosphoribosyltransferase (UPRT) from the protozoan Toxoplasma gondii. T. gondii UPRT has been well characterized because of its application in monitoring parasite growth: mammals lack this enzyme activity and thus only the parasite incorporates (3)H-uracil into its nucleic acids. In this study we used RNA labeling by UPRT to determine the roles of mRNA synthesis and decay in the control of gene expression during T. gondii asexual development. We also used this approach to specifically label parasite RNA during a mouse infection and to incorporate thio-substituted uridines into the RNA of human cells engineered to express T. gondii UPRT, indicating that engineered UPRT expression will allow cell-specific analysis of gene expression in organisms other than T. gondii.
Subject headings: Animals; Gene Expression Profiling/methods; Gene Expression Regulation/physiology; Humans; Metabolic Clearance Rate; Oligonucleotide Array Sequence Analysis/methods; Pentosyltransferases/chemistry/metabolism; RNA, Messenger/chemistry/genetics/metabolism; Signal Transduction/physiology; Staining and Labeling/methods; Toxoplasma/genetics/metabolism; Transcription Factors/metabolism; Transcriptional Activation/physiology
Publication year: 2005
Journal or book title: Nature Biotechnology
Volume: 23
Issue: 2
Pages: 232-237
Find the full text : https://www.nature.com/articles/nbt1061
Find more like this one (cited by): https://scholar.google.com/scholar?cites=13737379570745922183&as_sdt=1000005&sciodt=0,16&hl=en
Type: Journal Article
Serial number: 1344