Nucleotide Limitation Results inImpaired Photosynthesis, Reduced Growth andSeed Yield Together with Massively Altered Gene Expression.

Abstract

Nucleotide limitation and imbalance is a well-described phenomenon in animal research but understudied in the plant field. A peculiarity of pyrimidine de novo synthesis in plants is the complex subcellular organization. Here, we studied two organellar localized enzymes in the pathway, with chloroplast aspartate transcarbamoylase (ATC) and mitochondrial dihydroorotate dehydrogenase (DHODH). ATC knock-downs were most severely affected, exhibiting low levels of pyrimidine nucleotides, a low energy state, reduced photosynthetic capacity and accumulation of reactive oxygen species. Furthermore, altered leaf morphology and chloroplast ultrastructure were observed in ATC mutants. Although less affected, DHODH knock-down mutants showed impaired seed germination and altered mitochondrial ultrastructure. Thus, DHODH might not only be regulated by respiration but also exert a regulatory function on this process. Transcriptome analysis of an ATC-amiRNA line revealed massive alterations in gene expression with central metabolic pathways being downregulated and stress response and RNA-related pathways being upregulated. In addition, genes involved in central carbon metabolism, intracellular transport and respiration were markedly downregulated in ATC mutants, being most likely responsible for the observed impaired growth. We conclude that impairment of the first committed step in pyrimidine metabolism, catalyzed by ATC, leads to nucleotide limitation and by this has far-reaching consequences on metabolism and gene expression. DHODH might closely interact with mitochondrial respiration, as seen in delayed germination, which is the reason for its localization in this organelle.