Switching cell fate by the actin–auxin oscillator in Taxus: cellular aspects of plant cell fermentation

Plant cell fermentation, the production of valuable products in plant cell culture, has great potential as sustainable alternative to the exploitation of natural resources for compounds of pharmaceutical interest. However, the success of this approach has remained limited, because the cellular aspects of metabolic competence are mostly unknown. The production of the anti-cancer alkaloid Paclitaxel has been, so far, the most successful case for this approach. In the current work, we map cellular aspects of alkaloid synthesis in cells of Taxus chinensis using a combination of live-cell imaging, quantitative physiology, and metabolite analysis. We show evidence that metabolic potency correlates with a differentiation event giving rise to cells with large vacuoles with a tonoplast that is of a glossy appearance, agglomerations of lipophilic compounds, and multivesicular bodies that fuse with the plasma membrane. Cellular features of these glossy cells are bundled actin, more numerous peroxisomes, and vermiform mitochondria. The incidence of glossy cells can be increased by aluminium ions, and this increase is significantly reduced by the actin inhibitor Latrunculin B, and by diphenylene iodonium, a specific inhibitor of the NADPH oxidase Respiratory burst oxidase Homologue (RboH). It is also reduced by the artificial auxin Picloram. This cellular fingerprint matches the implications of a model, where the differentiation into the glossy cell type is regulated by the actin–auxin oscillator that in plant cells acts as dynamic switch between growth and defence

Cell type matters: competence for alkaloid metabolism differs in two seed-derived cell strains of Catharanthus roseus

Since the discovery of the anticancer drugs vinblastine and vincristine, Catharanthus roseus has been intensively studied for biosynthesis of several terpene indole alkaloids (TIAs). Due to their low abundance in plant tissues at a simultaneously high demand, modes of production alternative to conventional extraction are mandatory. Plant cell fermentation might become one of these alternatives, yet decades of research have shown limited success to certain product classes, leading to the question: how to preserve the intrinsic ability to produce TIAs (metabolic competence) in cell culture? We used the strategy to use the developmental potency of mature embryos to generate such strains. Two cell strains (C1and C4) from seed embryos of Catharanthus roseus were found to differ not only morphologically, but also in their metabolic competence. This differential competence became manifest not only under phytohormone elicitation, but also upon feeding with alkaloid pathway precursors. The more active strain C4 formed larger cell aggregates and was endowed with longer mitochondria. These cellular features were accompanied by higher alkaloid accumulation in response to methyl jasmonate (MeJA) elicitation. The levels of catharanthine could be increased significantly, while the concurrent vindoline branch of the pathway was blocked, such that no bisindole alkaloids were detectable. By feeding vindoline to MeJA-elicited C4 cells, vincristine became detectable; however, only to marginal amounts. In conclusion, these results show that cultured cells are not “de-differentiated”, but can differ in metabolic competence. In addition to elicitation and precursor feeding, the cellular properties of the “biomatter” are highly relevant for the success of plant cell fermentation

Gain time to adapt: How sorghum acquires tolerance to salinity

Salinity is a global environmental threat to agricultural production and food security around the world. To delineate salt-induced damage from adaption events we analysed a pair of sorghum genotypes which are contrasting in their response to salt stress with respect to physiological, cellular, metabolomic, and transcriptional responses. We find that the salt-tolerant genotype Della can delay the transfer of sodium from the root to the shoot, more swiftly deploy accumulation of proline and antioxidants in the leaves and transfer more sucrose to the root as compared to its susceptible counterpart Razinieh. Instead Razinieh shows metabolic indicators for a higher extent photorespiration under salt stress. Following sodium accumulation by a fluorescent dye in the different regions of the root, we find that Della can sequester sodium in the vacuoles of the distal elongation zone. The timing of the adaptive responses in Della leaves indicates a rapid systemic signal from the roots that is travelling faster than sodium itself. We arrive at a model where resistance and susceptibility are mainly a matter of temporal patterns in signalling

Physiological and Proteomic Analysis of the Rice Mutant cpm2 Suggests a Negative Regulatory Role of Jasmonic Acid in Drought Tolerance

It is widely known that numerous adaptive responses of drought-stressed plants are stimulated by chemical messengers known as phytohormones. Jasmonic acid (JA) is one such phytohormone. But there are very few reports revealing its direct implication in drought related responses or its cross-talk with other phytohormones. In this study, we compared the morpho-physiological traits and the root proteome of a wild type (WT) rice plant with its JA biosynthesis mutant coleoptile photomorphogenesis 2 (cpm2), disrupted in the allene oxide cyclase (AOC) gene, for insights into the role of JA under drought. The mutant had higher stomatal conductance, higher water use efficiency and higher shoot ABA levels under severe drought as compared to the WT. Notably, roots of cpm2 were better developed compared to the WT under both, control and drought stress conditions. Root proteome was analyzed using the Tandem Mass Tag strategy to better understand this difference at the molecular level. Expectedly, AOC was unique but notably highly abundant under drought in the WT. Identification of other differentially abundant proteins (DAPs) suggested increased energy metabolism (i.e., increased mobilization of resources) and reactive oxygen species scavenging in cpm2 under drought. Additionally, various proteins involved in secondary metabolism, cell growth and cell wall synthesis were also more abundant in cpm2 roots. Proteome-guided transcript, metabolite, and histological analyses provided further insights into the favorable adaptations and responses, most likely orchestrated by the lack of JA, in the cpm2 roots. Our results in cpm2 are discussed in the light of JA crosstalk to other phytohormones. These results together pave the path for understanding the precise role of JA during drought stress in rice

Physiological and Proteomic Analysis of the Rice Mutant cpm2 Suggests a Negative Regulatory Role of Jasmonic Acid in Drought Tolerance

It is widely known that numerous adaptive responses of drought-stressed plants are stimulated by chemical messengers known as phytohormones. Jasmonic acid (JA) is one such phytohormone. But there are very few reports revealing its direct implication in drought related responses or its cross-talk with other phytohormones. In this study, we compared the morpho-physiological traits and the root proteome of a wild type (WT) rice plant with its JA biosynthesis mutant coleoptile photomorphogenesis 2 (cpm2), disrupted in the allene oxide cyclase (AOC) gene, for insights into the role of JA under drought. The mutant had higher stomatal conductance, higher water use efficiency and higher shoot ABA levels under severe drought as compared to the WT. Notably, roots of cpm2 were better developed compared to the WT under both, control and drought stress conditions. Root proteome was analyzed using the Tandem Mass Tag strategy to better understand this difference at the molecular level. Expectedly, AOC was unique but notably highly abundant under drought in the WT. Identification of other differentially abundant proteins (DAPs) suggested increased energy metabolism (i.e., increased mobilization of resources) and reactive oxygen species scavenging in cpm2 under drought. Additionally, various proteins involved in secondary metabolism, cell growth and cell wall synthesis were also more abundant in cpm2 roots. Proteome-guided transcript, metabolite, and histological analyses provided further insights into the favorable adaptations and responses, most likely orchestrated by the lack of JA, in the cpm2 roots. Our results in cpm2 are discussed in the light of JA crosstalk to other phytohormones. These results together pave the path for understanding the precise role of JA during drought stress in rice

Genome-wide association study on metabolite accumulation in a wild barley NAM population reveals natural variation in sugar metabolism.

Metabolites play a key role in plants as they are routing plant developmental processes and are involved in biotic and abiotic stress responses. Their analysis can offer important information on the underlying processes. Regarding plant breeding, metabolite concentrations can be used as biomarkers instead of or in addition to genetic markers to predict important phenotypic traits (metabolic prediction). In this study, we applied a genome-wide association study (GWAS) in a wild barley nested association mapping (NAM) population to identify metabolic quantitative trait loci (mQTL). A set of approximately 130 metabolites, measured at early and late sampling dates, was analysed. For four metabolites from the early and six metabolites from the late sampling date significant mQTL (grouped as 19 mQTL for the early and 25 mQTL for the late sampling date) were found. Interestingly, all of those metabolites could be classified as sugars. Sugars are known to be involved in signalling, plant growth and plant development. Sugar-related genes, encoding mainly sugar transporters, have been identified as candidate genes for most of the mQTL. Moreover, several of them co-localized with known flowering time genes like Ppd-H1, HvELF3, Vrn-H1, Vrn-H2 and Vrn-H3, hinting on the known role of sugars in flowering. Furthermore, numerous disease resistance-related genes were detected, pointing to the signalling function of sugars in plant resistance. An mQTL on chromosome 1H in the region of 13 Mbp to 20 Mbp stood out, that alone explained up to 65% of the phenotypic variation of a single metabolite. Analysis of family-specific effects within the diverse NAM population showed the available natural genetic variation regarding sugar metabolites due to different wild alleles. The study represents a step towards a better understanding of the genetic components of metabolite accumulation, especially sugars, thereby linking them to biological functions in barley

Can metabolic prediction be an alternative to genomic prediction in barley?

Like other crop species, barley, the fourth most important crop worldwide, suffers from the genetic bottleneck effect, where further improvements in performance through classical breeding methods become difficult. Therefore, indirect selection methods are of great interest. Here, genomic prediction (GP) based on 33,005 SNP markers and, alternatively, metabolic prediction (MP) based on 128 metabolites with sampling at two different time points in one year, were applied to predict multi-year agronomic traits in the nested association mapping (NAM) population HEB-25. We found prediction abilities of up to 0.93 for plant height with SNP markers and of up to 0.61 for flowering time with metabolites. Interestingly, prediction abilities in GP increased after reducing the number of incorporated SNP markers. The estimated effects of GP and MP were highly concordant, indicating MP as an interesting alternative to GP, being able to reflect a stable genotype-specific metabolite profile. In MP, sampling at an early developmental stage outperformed sampling at a later stage. The results confirm the value of GP for future breeding. With MP, an interesting alternative was also applied successfully. However, based on our results, usage of MP alone cannot be recommended in barley. Nevertheless, MP can assist in unravelling physiological pathways for the expression of agronomically important traits

Presentation1.pdf

<p>It is widely known that numerous adaptive responses of drought-stressed plants are stimulated by chemical messengers known as phytohormones. Jasmonic acid (JA) is one such phytohormone. But there are very few reports revealing its direct implication in drought related responses or its cross-talk with other phytohormones. In this study, we compared the morpho-physiological traits and the root proteome of a wild type (WT) rice plant with its JA biosynthesis mutant coleoptile photomorphogenesis 2 (cpm2), disrupted in the allene oxide cyclase (AOC) gene, for insights into the role of JA under drought. The mutant had higher stomatal conductance, higher water use efficiency and higher shoot ABA levels under severe drought as compared to the WT. Notably, roots of cpm2 were better developed compared to the WT under both, control and drought stress conditions. Root proteome was analyzed using the Tandem Mass Tag strategy to better understand this difference at the molecular level. Expectedly, AOC was unique but notably highly abundant under drought in the WT. Identification of other differentially abundant proteins (DAPs) suggested increased energy metabolism (i.e., increased mobilization of resources) and reactive oxygen species scavenging in cpm2 under drought. Additionally, various proteins involved in secondary metabolism, cell growth and cell wall synthesis were also more abundant in cpm2 roots. Proteome-guided transcript, metabolite, and histological analyses provided further insights into the favorable adaptations and responses, most likely orchestrated by the lack of JA, in the cpm2 roots. Our results in cpm2 are discussed in the light of JA crosstalk to other phytohormones. These results together pave the path for understanding the precise role of JA during drought stress in rice.</p