De novo pyrimidine biosynthesis in the oomycete plant pathogen Phytophthora infestans

Abstract

The oomycete Phytophthora infestans, causal agent of the tomato and potato late blight, generates important economic and environmental losses worldwide. As current control strategies are becoming less effective, there is a need for studies on oomycete metabolism to help identify promising and more effective targets for chemical control. The pyrimidine pathways are attractive metabolic targets to combat tumors, virus and parasitic diseases but have not yet been studied in Phytophthora. Pyrimidines are involved in several critical cellular processes and play structural, metabolic and regulatory functions. Here, we used genomic and transcriptomic information to survey the pyrimidine metabolism during the P. infestans life cycle. After assessing the putative gene machinery for pyrimidine salvage and de novo synthesis, we inferred genealogies for each enzymatic domain in the latter pathway, which displayed a mosaic origin. The last two enzymes of the pathway, orotate phosphoribosyltransferase and orotidine-5-monophosphate decarboxylase, are fused in a multi-domain enzyme and are duplicated in some P. infestans strains. Two splice variants of the third gene (dihydroorotase) were identified, one of them encoding a premature stop codon generating a non-functional truncated protein. Relative expression profiles of pyrimidine biosynthesis genes were evaluated by qRT-PCR during infection in Solanum phureja. The third and fifth genes involved in this pathway showed high up-regulation during biotrophic stages and down-regulation during necrotrophy, whereas the uracil phosphoribosyl transferase gene involved in pyrimidine salvage showed the inverse behavior. These findings suggest the importance of de novo pyrimidine biosynthesis during the fast replicative early infection stages and highlight the dynamics of the metabolism associated with the hemibiotrophic life style of pathogen.

Keywords: 5-phosphoribosyl-1-pyrophosphate; ATCase; Alternative splicing;; CPSaseII; DHODase; DHOase; Horizontal gene transfer;; ODCase; OPRTase; ORF; Oomycetes; PRPP; Phytophthora;; Pyrimidine metabolism;; UMP; UMPase; UPRTase; aspartate transcarbamylase; carbamoyl phosphate synthetase II; dihydroorotase; dihydroorotate dehydrogenase; hours post inoculation; hpi; open reading frame; orotate phosphoribosyltransferase; orotidine-5-monophosphate decarboxylase; qRT-PCR; quantitative reverse transcription PCR; uracil phosphoribosyltransferase; uridine monophosphate; uridine monophosphate synthetase.