Analysis of Pyrimidine Catabolism inDrosophila melanogasterUsing Epistatic Interactions With Mutations of Pyrimidine Biosynthesis and β-Alanine Metabolism

Abstract

The biochemical pathway for pyrimidine catabolism links the pathways for pyrimidine biosynthesis and salvage with b-alanine metabolism, providing an array of epistatic interactions with which to analyze mutations of these pathways. Loss-of-function mutations have been identified and characterized for each of the enzymes for pyrimidine catabolism: dihydropyrimidine dehydrogenase (DPD), su(r) mutants; dihydropyrimidinase (DHP), CRMP mutants; b-alanine synthase (bAS), pyd3 mutants. For all three genes, mutants are viable and fertile and manifest no obvious phenotypes, aside from a variety of epistatic interactions. Mutations of all three genes disrupt suppression by the rudimentary gain-of-function mutation (r Su(b) ) of the dark cuticle phenotype of black mutants in which b-alanine pools are diminished; these results confirm that pyrimidines are the major source of b-alanine in cuticle pigmentation. The truncated wing phenotype of rudimentary mutants is suppressed completely by su(r) mutations and partially by CRMP mutations; however, no suppression is exhibited by pyd3 mutations. Similarly, su(r) mutants are hypersensitive to dietary 5-fluorouracil, CRMP mutants are less sensitive, and pyd3 mutants exhibit wild-type sensitivity. These results are discussed in the context of similar consequences of 5-fluoropyrimidine toxicity and pyrimidine catabolism mutations in humans.

Systematic interaction network filtering identifies CRMP1 as a novel suppressor of huntingtin misfolding and neurotoxicity

“…The missense mutation D408V suppresses the effect of CRMP1 on mutant HTT aggregation and toxicity in model systems Previous studies in Drosophila indicate that an exchange of an aspartic acid at position 430 for a valine residue alters the function of CRMP (Rawls 2006), suggesting that exchanging the corresponding amino acid in human CRMP1 might also influence its function. We found that the aspartic acid at position 408 is highly conserved in human CRMP1 (Fig.…”

Divergent Functions Through Alternative Splicing: The Drosophila CRMP Gene in Pyrimidine Metabolism, Brain, and Behavior

“…For this study, several derivatives were created of the P{PYD2+} transgene that contains a 10.9-kb wild-type genomic DNA fragment spanning the CRMP gene (Figure 1) inserted in the pCaSpeR4 vector (Rawls 2006). P{PYD2GFP}…”

“…CRMP supA4 is a nonconservative missense mutation that abolishes DHP activity (Rawls 2006). The CRMP supK1 and CRMP supIa1 mutations are intragenic deletions of the CRMP gene and are apparent null mutations; origins and properties of both mutations are described in Supporting Information, File S1).…”

“…The P{EP}Rac2 EP3118 strain, obtained from the Szeged Drosophila Stock Center, contains the P{EPgy2} enhancer trap transposon inserted into the 59 end of the Rac2 gene, providing transcriptional control of Rac2 by GAL4-responsive elements within the transposon (Rørth et al 1998;Tseng and Hariharan 2002). The wildtype flies utilized in behavior experiments were Canton-S w 1118 (iso CJ1) from the Dubnau lab; all mutations and transgenes analyzed in the behavioral experiments were introduced into the iso CJ1 genetic background by backcrossing at least six generations.For this study, several derivatives were created of the P{PYD2+} transgene that contains a 10.9-kb wild-type genomic DNA fragment spanning the CRMP gene (Figure 1) inserted in the pCaSpeR4 vector (Rawls 2006). P{PYD2GFP}…”

“…The precise mechanism by which CRMP functions in PlexA/MICAL signaling is unknown. In Drosophila melanogaster, the CRMP gene is the sole gene for the DHP/CRMP protein family and has been shown to encode DHP (Rawls 2006). To better understand DHP/CRMP proteins in D. melanogaster, we have carried out experiments to further characterize the fly CRMP gene and its products.…”