Summary: Roots, tubers, and bananas (RTB) are vital staples for food security in the world's poorest nations. A major constraint to current RTB breeding programmes is limited knowledge on the available diversity due to lack of efficient germplasm characterization and structure. In recent years large‐scale efforts have begun to elucidate the genetic and phenotypic diversity of germplasm collections and populations and, yet, biochemical measurements have often been overlooked despite metabolite composition being directly associated with agronomic and consumer traits. Here we present a compound database and concentration range for metabolites detected in the major RTB crops: banana (Musa spp.), cassava (Manihot esculenta), potato (Solanum tuberosum), sweet potato (Ipomoea batatas), and yam (Dioscorea spp.), following metabolomics‐based diversity screening of global collections held within the CGIAR institutes. The dataset including 711 chemical features provides a valuable resource regarding the comparative biochemical composition of each RTB crop and highlights the potential diversity available for incorporation into crop improvement programmes. Particularly, the tropical crops cassava, sweet potato and banana displayed more complex compositional metabolite profiles with representations of up to 22 chemical classes (unknowns excluded) than that of potato, for which only metabolites from 10 chemical classes were detected. Additionally, over 20% of biochemical signatures remained unidentified for every crop analyzed. Integration of metabolomics with the on‐going genomic and phenotypic studies will enhance ’omics‐wide associations of molecular signatures with agronomic and consumer traits via easily quantifiable biochemical markers to aid gene discovery and functional characterization.

Cassava (Manihot esculenta Crantz) is the predominant staple food in Sub-Saharan African (SSA) and an industrial crop in South East Asia. Despite focused breeding efforts for increased yield, resistance and nutritional value, cassava breeding has not advanced at the same rapidity as other staple crops. In the present study, metabolomic techniques were developed and implemented to characterize the chemotypes of selected cassava accessions and assess potential resources for the breeding program. The metabolite data analysed was applied to describe the biochemical diversity available in the panel, identifying South American accessions as the most diverse. Genotypes with distinct phenotypic traits showed a representative metabolite profile and could be clearly identified, even if the phenotypic trait was a root characteristic e.g. high amylose content.

Summary: The production of complex multimeric secretory immunoglobulins (SIgA) in Nicotiana benthamiana leaves is challenging, with significant reductions in complete protein assembly and consequently yield, being the most important difficulties. Expanding the physical dimensions of the ER to mimic professional antibody‐secreting cells can help to increase yields and promote protein folding and assembly. Here, we expanded the ER in N. benthamiana leaves by targeting the enzyme CTP:phosphocholine cytidylyltransferase (CCT), which catalyses the rate‐limiting step in the synthesis of the key membrane component phosphatidylcholine (PC). We used CRISPR/Cas to perform site‐directed mutagenesis of each of the three endogenous CCT genes in N. benthamiana by introducing frame‐shifting indels to remove the auto‐inhibitory C‐terminal domains. We generated stable homozygous lines of N. benthamiana containing different combinations of the edited genes, including plants where all three isofunctional CCT homologues were modified. Changes in ER morphology in the mutant plants were confirmed by in vivo confocal imaging and substantially increased the yields of two fully assembled SIgAs by prolonging the ER residence time and boosting chaperone accumulation. Through a combination of ER engineering with chaperone overexpression, we increased the yields of fully assembled SIgA by an order of magnitude, reaching almost 1 g/kg fresh leaf weight. This strategy removes a major roadblock to producing SIgA and will likely facilitate the production of other complex multimeric biopharmaceutical proteins in plants.

SUMMARY: Sustainable production of chemicals and improving these biosources by engineering metabolic pathways to create efficient plant‐based biofactories relies on the knowledge of available chemical/biosynthetic diversity present in the plant. Nicotiana species are well known for their amenability towards transformation and other new plant breeding techniques. The genus Nicotiana is primarily known through Nicotiana tabacum L., the source of tobacco leaves and all respective tobacco products. Due to the prevalence of the latter, N. tabacum and related Nicotiana species are one of the most extensively studied plants. The majority of studies focused solely on N. tabacum or other individual species for chemotyping. The present study analysed a diversity panel including 17 Nicotiana species and six accessions of Nicotiana benthamiana and created a data set that effectively represents the chemotype core collection of the genus Nicotiana. The utilisation of several analytical platforms and previously published libraries/databases enabled the identification and measurement of over 360 metabolites of a wide range of chemical classes as well as thousands of unknowns with dedicated spectral and chromatographic properties.

Light is an essential regulator of many developmental processes in higher plants. We investigated the effect of 4-hydroxy-3-methylbut-2-enyl diphosphate reductase 1/2 genes (OsHDR1/2) and isopentenyl diphosphate isomerase 1/2 genes (OsIPPI1/2) on the biosynthesis of chlorophylls, carotenoids, and phytosterols in 14-day-old etiolated rice (Oyza sativa L.) leaves during de-etiolation. However, little is known about the effect of isoprenoid biosynthesis genes on the corresponding metabolites during the de-etiolation of etiolated rice leaves. The results showed that the levels of α-tocopherol were significantly increased in de-etiolated rice leaves. Similar to 1-deoxy-D-xylulose-5-phosphate synthase 3 gene (OsDXS3), both OsDXS1 and OsDXS2 genes encode functional 1-deoxy-D-xylulose-5-phosphate synthase (DXS) activities. Their expression patterns and the synthesis of chlorophyll, carotenoid, and tocopherol metabolites suggested that OsDXS1 is responsible for the biosynthesis of plastidial isoprenoids in de-etiolated rice leaves. The expression analysis of isoprenoid biosynthesis genes revealed that the coordinated expression of the MEP (2-C-methyl-D-erythritol 4-phosphate) pathway, chlorophyll, carotenoid, and tocopherol pathway genes mirrored the changes in the levels of the corresponding metabolites during de-etiolation. The underpinning mechanistic basis of coordinated light-upregulated gene expression was elucidated during the de-etiolation process, specifically the role of light-responsive cis-regulatory motifs in the promoter region of these genes. In silico promoter analysis showed that the light-responsive cis-regulatory elements presented in all the promoter regions of each light-upregulated gene, providing an important link between observed phenotype during de-etiolation and the molecular machinery controlling expression of these genes.