Nucleotide Imbalance, Provoked by Downregulation of Aspartate Transcarbamoylase Impairs Cold Acclimation in Arabidopsis

Abstract

Aspartate transcarbamoylase (ATC) catalyzes the first committed step in pyrimidine de novo synthesis. As shown before, mutants with 80% reduced transcript and protein levels exhibit reduced levels of pyrimidine metabolites and thus nucleotide limitation and imbalance. Consequently, reduced photosynthetic capacity and growth, accompanied by massive transcriptional changes, were observed. Here, we show that nucleotide de novo synthesis was upregulated during cold acclimation of Arabidopsis thaliana (ecotype Columbia, Col-0) plants, but ATC knockdown mutants failed to acclimate to this condition as they did not accumulate neutral sugars and anthocyanins. A global transcriptome analysis revealed that most of the transcriptional changes observed in Col-0 plants upon cold exposure were also evident in ATC knockdown plants. However, several responses observed in cold-treated Col-0 plants could already be detected in knockdown plants when grown under standard conditions, suggesting that these mutants exhibited typical cold responses without prior cold stimulation. We believe that nucleotide signaling is involved in “cold-like priming” and “cold acclimation” in general. The observed transcript levels of genes involved in central carbon metabolism and respiration were an exception to these findings. These were upregulated in the cold but downregulated in warm-grown ATC mutants.

Transcriptional reprogramming of nucleotide metabolism in response to altered pyrimidine availability in Arabidopsis seedlings

“…Induction of pyrimidine catabolism genes, under conditions of severe pyrimidine limitation, was unexpected in PALA-treated seedlings. In two previous studies with ATC knockdown lines with less severe phenotypes (Chen and Slocum, 2008;Bellin et al, 2023a), catabolic pathway genes were repressed. In the present study, enhanced catabolism of labeled Ura in day 9 seedlings may have resulted from repression of Ura salvaging pathway genes, leading to Ura accumulation in the plastid and increased availability to PYD1 and the catabolic pathway.…”

“…For example, antisense-mediated knockdown of UMP synthase expression in the de novo pyrimidine synthesis pathway in potato tubers resulted in increased expression of genes involved in pyrimidine salvaging, increased salvaging pathway activities, and increased levels of uridine nucleotides as a compensatory response (Geigenberger et al, 2005). Similar results were obtained for Arabidopsis RNAi lines with decreased expression of aspartate transcarbamoylase (Chen and Slocum, 2008), which regulates de novo pyrimidine pathway activities (Schröder et al, 2005;Bellin et al, 2021;Bellin et al, 2023a). Increased expression of pyrimidine salvaging pathway genes and decreased expression of catabolic pathway genes in response to pyrimidine limitation suggested similar compensatory responses that could help maintain pyrimidine homeostasis in these plants.…”

UPP affects chloroplast development by interfering with chloroplast proteostasis

“…(PAM) fluorometry, as previously described (Bellin et al, 2023). Five-week-old plants grown under standard conditions were transferred to high light, and leaf discs from respective plants were treated with DCMU (5µM).…”

“…Photosynthetic activity was measured as described previously (Bellin et al, 2023). Five-week-old plants grown under standard conditions were transferred to high light.…”

“…Thereby, the de novo synthesis and catabolism of the two molecule classes, the purines and the pyrimidines, are mostly separated in terms of the enzymes involved (Henderson and Paterson, 1973;(Zrenner et al, 2006;Witte and Herde, 2020). However, to allow for proper nucleic acid synthesis, purines and pyrimidines need to be balanced (Bellin et al, 2023). This is achieved by transcription factors, allosteric regulation and feedback inhibition (Lane and Fan, 2015;.…”