Effects of impaired steryl ester biosynthesis on tomato growth and developmental processes.

Steryl esters (SE) are stored in cytoplasmic lipid droplets and serve as a reservoir of sterols that helps to maintain free sterols (FS) homeostasis in cell membranes throughout plant growth and development, and provides the FS needed to meet the high demand of these key plasma membrane components during rapid plant organ growth and expansion. SE are also involved in the recycling of sterols and fatty acids released from membranes during plant tissues senescence. SE are synthesized by sterol acyltransferases, which catalyze the transfer of long-chain fatty acid groups to the hydroxyl group at C3 position of FS. Depending on the donor substrate, these enzymes are called acyl-CoA:sterol acyltransferases (ASAT), when the substrate is a long-chain acyl-CoA, and phospholipid:sterol acyltransferases (PSAT), which use a phospholipid as a donor substrate. We have recently identified and preliminary characterized the tomato (Solanum lycopersicum cv. Micro-Tom) SlASAT1 and SlPSAT1 enzymes. To gain further insight into the biological role of these enzymes and SE biosynthesis in tomato, we generated and characterized CRISPR/Cas9 single knock-out mutants lacking SlPSAT1 (slpsat1) and SlASAT1 (slasat1), as well as the double mutant slpsat1 x slasat1. Analysis of FS and SE profiles in seeds and leaves of the single and double mutants revealed a strong depletion of SE in slpsat1, that was even more pronounced in the slpsat1 x slasat1 mutant, while an increase of SE levels was observed in slasat1. Moreover, SlPSAT1 and SlASAT1 inactivation affected in different ways several important cellular and physiological processes, like leaf lipid bo1dies formation, seed germination speed, leaf senescence, and the plant size. Altogether, our results indicate that SlPSAT1 has a predominant role in tomato SE biosynthesis while SlASAT1 would mainly regulate the flux of the sterol pathway. It is also worth to mention that some of the metabolic and physiological responses in the tomato mutants lacking functional SlPSAT1 or SlASAT1 are different from those previously reported in Arabidopsis, being remarkable the synergistic effect of SlASAT1 inactivation in the absence of a functional SlPSAT1 on the early germination and premature senescence phenotypes.

The Emergence of Genome Editing-Innovation Network Dynamics of Academic Publications, Patents, and Business Activities.

Transformative societal change can both be triggered and influenced by both macro-level political means and the emergence of technologies. Key enabling technologies and therein biotechnology hold the power to drive those changes forward, evolving from breakthrough academic discoveries into business activities. Due to its increasing empirical relevance, we picked genome editing as an example for an emerging technology and extracted publication, patent, and company data from the years 2000 to 2020. By drawing upon social network analysis, we identify major networks and clusters that are dominating the respective time and layer. Based on these networks, we draw vertical connections between scientific knowledge, patented technologies, and business activities to visualize the interlevel relationships between actors through technological development. Thereby, we identify network dynamics of the emergence of genome editing, the most important actors and clusters evolving, and its spread into different areas.

Genome Editing in Crop Plant Research-Alignment of Expectations and Current Developments.

The rapid development of genome editing and other new genomic techniques (NGT) has evoked manifold expectations on purposes of the application of these techniques to crop plants. In this study, we identify and align these expectations with current scientific development. We apply a semi-quantitative text analysis approach on political, economic, and scientific opinion papers to disentangle and extract expectations towards the application of NGT-based plants. Using the sustainable development goals (SDG) of the 2030 agenda as categories, we identify contributions to food security or adaptation to climatic changes as the most frequently mentioned expectations, accompanied by the notion of sustainable agriculture and food systems. We then link SDG with relevant plant traits and review existing research and commercial field trials for genome-edited crop plants. For a detailed analysis we pick as representative traits drought tolerance and resistance against fungal pathogens. Diverse genetic setscrews for both traits have been identified, modified, and tested under laboratory conditions, although there are only a few in the field. All in all, NGT-plants that can withstand more than one stressor or different environments are not documented in advanced development states. We further conclude that developing new plants with modified traits will not be sufficient to reach food security or adaption to climatic changes in a short time frame. Further scientific development of sustainable agricultural systems will need to play an important role to tackle SDG challenges, as well.

An underutilized orphan tuber crop-Chinese yam : a review.

MAIN CONCLUSION: The diversification of food crops can improve our diets and address the effects of climate change, and in this context the orphan crop Chinese yam shows significant potential as a functional food. As the effects of climate change become increasingly visible even in temperate regions, there is an urgent need to diversify our crops in order to address hunger and malnutrition. This has led to the re-evaluation of neglected species such as Chinese yam (Dioscorea polystachya Turcz.), which has been cultivated for centuries in East Asia as a food crop and as a widely-used ingredient in traditional Chinese medicine. The tubers are rich in nutrients, but also contain bioactive metabolites such as resistant starches, steroidal sapogenins (like diosgenin), the storage protein dioscorin, and mucilage polysaccharides. These health-promoting products can help to prevent cardiovascular disease, diabetes, and disorders of the gut microbiome. Whereas most edible yams are tropical species, Chinese yam could be cultivated widely in Europe and other temperate regions to take advantage of its nutritional and bioactive properties. However, this is a laborious process and agronomic knowledge is fragmented. The underground tubers contain most of the starch, but are vulnerable to breaking and thus difficult to harvest. Breeding to improve tuber shape is complex given the dioecious nature of the species, the mostly vegetative reproduction via bulbils, and the presence of more than 100 chromosomes. Protocols have yet to be established for in vitro cultivation and genetic transformation, which limits the scope of research. This article summarizes the sparse research landscape and evaluates the nutritional and medical applications of Chinese yam. By highlighting the potential of Chinese yam tubers, we aim to encourage the adoption of this orphan crop as a novel functional food.

Taraxacum brevicorniculatum rubber elongation factor TbREF associates with lipid droplets and affects lipid turn-over in yeast.

A protein named TbREF that is localized on rubber particles of the rubber producing dandelion species Taraxacum brevicorniculatum was expressed in tobacco leaves and in yeast. TbREF fused to fluorescence proteins colocalized on globular, hydrophobic structures, most likely lipid droplets. Furthermore, triacylglycerol, sterol and total lipid content of TbREF expressing yeast was determined by photometric analyses of nile red stainings and GC-MS analyses. Therefore, yeast exposed an enhanced nile red fluorescence as well as an increased TAG and sterol content compared to wildtype and vector control. Altogether, these findings gave new insights into the putative function of TbREF that might be pushing rubber particle production due to its cytotoxic nature and/or shielding and preventing degradation of lipid droplets. Furthermore, these results highlight possible biotechnological applications regarding the accumulation of hydrophobic compounds in lipid droplet like structures.

Small rubber particle proteins from Taraxacum brevicorniculatum promote stress tolerance and influence the size and distribution of lipid droplets and artificial poly(cis-1,4-isoprene) bodies.

Natural rubber biosynthesis occurs on rubber particles, i.e. organelles resembling small lipid droplets localized in the laticifers of latex-containing plant species, such as Hevea brasiliensis and Taraxacum brevicorniculatum. The latter expresses five small rubber particle protein (SRPP) isoforms named TbSRPP1-5, the most abundant proteins in rubber particles. These proteins maintain particle stability and are therefore necessary for rubber biosynthesis. TbSRPP1-5 were transiently expressed in Nicotiana benthamiana protoplasts and the proteins were found to be localized on lipid droplets and in the endoplasmic reticulum, with TbSRPP1 and TbSRPP3 also present in the cytosol. Bimolecular fluorescence complementation confirmed pairwise interactions between all proteins except TbSRPP2. The corresponding genes showed diverse expression profiles in young T. brevicorniculatum plants exposed to abiotic stress, and all except TbSRPP4 and TbSRPP5 were upregulated. Young Arabidopsis thaliana plants that overexpressed TbSRPP2 and TbSRPP3 tolerated drought stress better than wild-type plants. Furthermore, we used rubber particle extracts and standards to investigate the affinity of the TbSRPPs for different phospholipids, revealing a preference for negatively charged head groups and 18:2/16:0 fatty acid chains. This finding may explain the effect of TbSRPP3-5 on the dispersity of artificial poly(cis-1,4-isoprene) bodies and on the lipid droplet distribution we observed in N. benthamiana leaves. Our data provide insight into the assembly of TbSRPPs on rubber particles, their role in rubber particle structure, and the link between rubber biosynthesis and lipid droplet-associated stress responses, suggesting that SRPPs form the basis of evolutionarily conserved intracellular complexes in plants.

Identification of a Taraxacum brevicorniculatum rubber elongation factor protein that is localized on rubber particles and promotes rubber biosynthesis.

Two protein families required for rubber biosynthesis in Taraxacum brevicorniculatum have recently been characterized, namely the cis-prenyltransferases (TbCPTs) and the small rubber particle proteins (TbSRPPs). The latter were shown to be the most abundant proteins on rubber particles, where rubber biosynthesis takes place. Here we identified a protein designated T. brevicorniculatum rubber elongation factor (TbREF) by using mass spectrometry to analyze rubber particle proteins. TbREF is homologous to the TbSRPPs but has a molecular mass that is atypical for the family. The promoter was shown to be active in laticifers, and the protein itself was localized on the rubber particle surface. In TbREF-silenced plants generated by RNA interference, the rubber content was significantly reduced, correlating with lower TbCPT protein levels and less TbCPT activity in the latex. However, the molecular mass of the rubber was not affected by TbREF silencing. The colloidal stability of rubber particles isolated from TbREF-silenced plants was also unchanged. This was not surprising because TbREF depletion did not affect the abundance of TbSRPPs, which are required for rubber particle stability. Our findings suggest that TbREF is an important component of the rubber biosynthesis machinery in T. brevicorniculatum, and may play a role in rubber particle biogenesis and influence rubber production.

The characteristics and potential applications of structural lipid droplet proteins in plants.

Plant cytosolic lipid droplets are storage organelles that accumulate hydrophobic molecules. They are found in many tissues and their general structure includes an outer lipid monolayer with integral and associated proteins surrounding a hydrophobic core. Two distinct types can be distinguished, which we define here as oleosin-based lipid droplets (OLDs) and non-oleosin-based lipid droplets (NOLDs). OLDs are the best characterized lipid droplets in plants. They are primarily restricted to seeds and other germinative tissues, their surface is covered with oleosin-family proteins to maintain stability, they store triacylglycerols (TAGs) and they are used as a source of energy (and possibly signaling molecules) during the germination of seeds and pollen. Less is known about NOLDs. They are more abundant than OLDs and are distributed in many tissues, they accumulate not only TAGs but also other hydrophobic molecules such as natural rubber, and the structural proteins that stabilize them are unrelated to oleosins. In many species these proteins are members of the rubber elongation factor superfamily. NOLDs are not typically used for energy storage but instead accumulate hydrophobic compounds required for environmental interactions such as pathogen defense. There are many potential applications of NOLDs including the engineering of lipid production in plants and the generation of artificial oil bodies.