Abstract
Biomass production requires the coordination between growth and metabolism. In a large-scale screen for mutants affected in leaf morphology, we isolated the orbiculata1 (orb1) mutants, which exhibit a pale green phenotype and reduced growth. The combination of map-based cloning and next-generation sequencing allowed us to establish that ORB1 encodes the GLUTAMATE SYNTHASE 1 (GLU1) enzyme, also known as FERREDOXIN-DEPENDENT GLUTAMINE OXOGLUTARATE AMINOTRANSFERASE 1 (Fd-GOGAT1). We performed an RNA-seq analysis to identify global gene expression changes in the orb1–3 mutant. We found altered expression levels of genes encoding enzymes involved in nitrogen assimilation and amino acid biosynthesis, such as glutamine synthetases, asparagine synthetases and glutamate dehydrogenases, showing that the expression of these genes depends on the levels of glutamine and/or glutamate. In addition, we observed a concerted upregulation of genes encoding subunits of the cytosolic ribosome. A gene ontology (GO) analysis of the differentially expressed genes between Ler and orb1–3 showed that the most enriched GO terms were ‘translation’, ‘cytosolic ribosome’ and ‘structural constituent of ribosome’. The upregulation of ribosome-related functions might reflect an attempt to keep protein synthesis at optimal levels even when the pool of glutamate is reduced.
Highlights
The final shape and size of leaves depends on a complex sequence of developmental events, which include the recruitment of cells to the incipient leaf primordium, the control of cell proliferation, the transition from cell proliferation to cell expansion, and cell expansion and differentiation[1,2,3]
We have found ORB1 to be the same gene as At5g04140, known as GLUTAMATE SYNTHASE 1 (GLU1) and FERREDOXIN-DEPENDENT GLUTAMINE OXOGLUTARATE AMINOTRANSFERASE 1 (Fd-GOGAT1)
Together with the conversion of glutamate and ammonium into glutamine using ATP, which is catalyzed by GLUTAMINE SYNTHETASE 2 (GS2), this reaction is an essential component of the GS/GOGAT cycle
Summary
The final shape and size of leaves depends on a complex sequence of developmental events, which include the recruitment of cells to the incipient leaf primordium, the control of cell proliferation, the transition from cell proliferation to cell expansion, and cell expansion and differentiation[1,2,3]. As an alternative approach, modifying the expression levels of key enzymes in primary carbon and nitrogen metabolic pathways might lead to increased biomass production[17]. The VENOSA3 (VEN3) and VEN6 genes of Arabidopsis encode the two subunits of the carbamoyl phosphate synthetase, which catalyzes the conversion of glutamine and bicarbonate into carbamoyl phosphate and glutamate in the arginine biosynthesis pathway[21]. Our RNA-seq profiling of orb[1] mutant leaves has given insight on how nitrogen-related metabolic pathways are regulated during the vegetative phase, and shows some differences with the results of a microarray analysis previously performed using a different allele of the same gene[33]. Our analysis of RNA-seq results has unveiled a concerted transcriptional increase of genes encoding components of the translational machinery in orb[1] mutants
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