UMP Pyrophosphorylase: A Moonlighting Protein with Essential Functions in Chloroplast Development and Photosynthesis Establishment

Abstract

Pyrimidine nucleotides (e.g. deoxy-CTP, deoxy-TTP, UTP, and CTP) are essential building blocks for DNA and RNA synthesis in all organisms. In plants, Suc and cell wall polymer synthesis depend on pyrimidine nucleotide-derived substrates, such as UDP-Glc, and important classes of membrane lipids are made using CDP-activated substrates. In addition to de novo synthesis pathways, pyrimidine nucleotides are produced via salvage pathways. Uracil is salvaged by uracil phosphoribosyl transferase (UPRT; EC 2.4.2.9), using phosphoribosyl pyrophosphate as substrate to produce UMP, while uridine is salvaged by uridine kinase (EC 2.7.1.48), which uses ATP to phosphorylate uridine to UMP. In this manuscript (Ohler et al., 2019), the authors show that pyrimidine salvage in Arabidopsis (Arabidopsis thaliana) occurs predominantly via uridine/ cytidine (Fig. 1), with the main enzymes being the bifunctional cytosolic uridine/cytidine kinases 1 and 2 (UCK1 and UCK2; EC 2.7.1.48). A double knockout of UCK1 and UCK2 leads to severe growth restrictions in green tissue (Chen and Thelen, 2011), indicating that Arabidopsis relies on both de novo pyrimidine synthesis and pyrimidine salvage via cytosolic uridine/ cytidine. These enzymes have a UPRT domain but do not exhibit any UPRT activity. The only enzyme responsible for UPRT activity in Arabidopsis is the chloroplastic UMP pyrophosphorylase (UPP; EC 2.4.2.9), which is involved in pyrimidine salvage via uracil, but despite being functional, this plastidic uracil salvage pathway has a nonessential role in Arabidopsis. Surprisingly, upp-1, a UPP T-DNA insertion line, germinates but dies at the seedling stage. This knockout exhibits severe growth defects, with pale yellow leaves, reduced chloroplast size and lack of starch synthesis, pointing to a role for UPP in chloroplast development

UPP affects chloroplast development by interfering with chloroplast proteostasis

“…This, is agrees with other results and suggests that an unknown second function is responsible for the inability of upp-1 to establish photosynthesis (Mainguet et al, 2009;Arrivault, 2019;Ohler et al, 2019).…”

“…Seedling lethal upp-1 can be rescued by complementation with nearly catalytic inactive UPP but not with a highly active homolog from Escherichia coli . This agrees with previous results and suggests that an unknown second (moonlighting) function is responsible for the inability of upp-1 to establish photosynthesis (Mainguet et al, 2009; Arrivault, 2019; Ohler et al, 2019).…”

“…UPP expression is high in the light and upp-1 grows like wild type in the dark (Ohler et al, 2019). Altogether it can be stated that UPP is required for the establishment of photosynthesis (Arrivault, 2019;. Artificial micro-RNA lines generated in the course of this work resemble the upp-1 phenotype in milder form with respect to photosynthetic efficiency and growth, for example (Figure 1, 6).…”

“…Additionally, downstream enzymes such as plastid UMP kinase (PUMPKIN) and nucleoside diphosphate kinase 2 (NDPK2) have also been found to exhibit moonlighting functions (Dorion and Rivoal, 2015;Arrivault, 2019;Schmid et al, 2019). Since knock-out mutants of both enzymes are not lethal to seedlings, as is the case with UPP, these moonlighting functions are likely distinct from those of UPP.…”

“…However, previous work provided evidence for a moonlighting function of Arabidopsis UPP of so far unknown nature (Ohler et al, 2019). Interestingly, the two downstream enzymes plastid UMP kinase (PUMPKIN) and nucleoside diphosphate kinase 2 (NDPK2) also exhibit moonlighting functions (Dorion and Rivoal, 2015;Arrivault, 2019;Schmid et al, 2019). Since knock-out mutants of both enzymes are not seedling lethal, as it is the case with UPP, these moonlighting functions are most likely distinct from those of UPP.…”