title: A link between impaired purine nucleotide synthesis and apoptosis in Drosophila melanogaster.
abstract: The biosynthetic pathways and multiple functions of purine nucleotides are well known. However, the pathways that respond to alterations in purine nucleotide synthesis in vivo in an animal model organism have not been identified. We examined the effects of inhibiting purine de novo synthesis in vivo and in cultured cells of Drosophila melanogaster. The purine de novo synthesis gene ade2 encodes phosphoribosylformylglycinamidine synthase (EC 6.3.5.3). An ade2 deletion, generated by P-element transposon excision, causes lethality in early pupal development, with darkening, or necrosis, of leg and wing imaginal disc tissue upon disc eversion. Together with analysis of a previously isolated weaker allele, ade2(4), and an allele of the Prat gene, which encodes an enzyme for the first step in the pathway, we determined that the lethal arrest and imaginal disc phenotypes involve apoptosis. A transgene expressing the baculovirus caspase inhibitor p35, which suppresses apoptosis caused by other stresses such as DNA damage, suppresses both the imaginal disc tissue darkening and the pupal lethality of all three purine de novo synthesis mutants. Furthermore, we showed the presence of apoptosis at the cellular level in both ade2 and Prat mutants by detecting TUNEL-positive nuclei in wing imaginal discs. Purine de novo synthesis inhibition was also examined in tissue culture by ade2 RNA interference followed by analysis of genome-wide changes in transcript levels. Among the upregulated genes was HtrA2, which encodes an apoptosis effector and is thus a candidate for initiating apoptosis in response to purine depletion.
authors: Department of Biology, University of New Brunswick, Fredericton, New Brunswick, Canada.
authors: Catherine
authors: C
authors: Holland
authors: David B
authors: DB
authors: Lipsett
authors: Denise V
authors: DV
authors: Clark
mesh_terms: D000582:Amidophosphoribosyltransferase / Q000235:genetics / Q000378:metabolism; D000818:Animals; D030801:Animals, Genetically Modified; D017209:Apoptosis; D001483:Base Sequence; D053898:Biosynthetic Pathways; D015153:Blotting, Western; D019731:Carbon-Nitrogen Ligases / Q000235:genetics / Q000378:metabolism; D019733:Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D003433:Crosses, Genetic; D004251:DNA Transposable Elements / Q000235:genetics; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster / Q000166:cytology / Q000235:genetics* / Q000378:metabolism*; D005260:Female; D020869:Gene Expression Profiling; D000074923:High-Temperature Requirement A Serine Peptidase 2; D020287:In Situ Nick-End Labeling; D008297:Male; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D011679:Pupa / Q000235:genetics / Q000378:metabolism; D011685:Purine Nucleotides / Q000096:biosynthesis*; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism
chemical_list: D004251:DNA Transposable Elements; D029721:Drosophila Proteins; D011685:Purine Nucleotides; D000582:Amidophosphoribosyltransferase; C531864:PRAT protein, Drosophila; D012697:Serine Endopeptidases; D000074923:High-Temperature Requirement A Serine Peptidase 2; C530280:HtrA2 protein, Drosophila; D019731:Carbon-Nitrogen Ligases; D019733:Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor; C561706:phosphoribosylformylglycinamidine synthase, Drosophila; C022385:phosphoribosylformylglycinamidine synthetase