Qualterra – Scalable biomass processing technologies for sustainable agriculture

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Involvement of G-proteins, calmodulin and tagetitoxin-sensitive RNA polymerase in light-regulated expression of plastid genes (psbA, psaA and rbcL) in rice (Oryza sativa L.)

The regulation of chloroplast gene expression by light involves multiple signaling components. In an earlier study, we demonstrated the role of calcium and phosphorylation in regulating the expression of photosynthesis-related plastid genes, psbA, psaA and rbcL, using rice as a model monocot system. This work has been extended further to examine the possible involvement of heterotrimeric GTP-binding proteins and calmodulin. Vacuum infiltration of 5-day-old etiolated rice seedlings with G-protein agonists, cholera toxin and GTPγS, increased the steady-state transcript levels of the plastid genes. The antagonists/inhibitors of calmodulin action, trifluoperazine and W7, inhibited the light-induced increase in steady-state transcript levels of these genes. The light-regulated expression of photosynthetic genes was also adversely affected by tagetitoxin, a specific inhibitor of plastid-encoded RNA polymerase. These results indicate the involvement of various signaling components in transduction of light signal that probably also recruits PEP to regulate plastid gene expression

Enhanced Translation of a Chloroplast-Expressed RbcS Gene Restores Small Subunit Levels and Photosynthesis in Nuclear RbcS Antisense Plants

Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is a key enzyme that converts atmospheric carbon to food and supports life on this planet. Its low catalytic activity and specificity for oxygen leads to photorespiration, severely limiting photosynthesis and crop productivity. Consequently, Rubisco is a primary target for genetic engineering. Separate localization of the genes in the nuclear and chloroplast genomes and a complex assembly process resulting in a very low catalytic activity of hybrid Rubisco enzymes have rendered several earlier attempts of Rubisco engineering unsuccessful. Here we demonstrate that the RbcS gene, when integrated at a transcriptionally active spacer region of the chloroplast genome, in a nuclear RbcS antisense line and expressed under the regulation of heterologous (gene 10) or native (psbA) UTRs, results in the assembly of a functional holoenzyme and normal plant growth under ambient CO2 conditions, fully shortcircuiting nuclear control of gene regulation. There was ≈150-fold more RbcS transcript in chloroplast transgenic lines when compared with the nuclear RbcS antisense line, whereas the wild type has 7-fold more transcript. The small subunit protein levels in the gene 10/RbcS and psbA/RbcS plants were 60% and 106%, respectively, of the wild type. Photosynthesis of gene 10/RbcS plants was approximately double that of the antisense plants, whereas that of psbA/RbcS plants was restored almost completely to the wild-type rates. These results have opened an avenue for using chloroplast engineering for the evaluation of foreign Rubisco genes in planta that eventually can result in achieving efficient photosynthesis and increased crop productivity

Comparative chloroplast genomics and phylogenetics of Fagopyrum esculentum ssp. ancestrale – A wild ancestor of cultivated buckwheat

<p>Abstract</p> <p>Background</p> <p>Chloroplast genome sequences are extremely informative about species-interrelationships owing to its non-meiotic and often uniparental inheritance over generations. The subject of our study, <it>Fagopyrum esculentum</it>, is a member of the family Polygonaceae belonging to the order Caryophyllales. An uncertainty remains regarding the affinity of Caryophyllales and the asterids that could be due to undersampling of the taxa. With that background, having access to the complete chloroplast genome sequence for <it>Fagopyrum </it>becomes quite pertinent.</p> <p>Results</p> <p>We report the complete chloroplast genome sequence of a wild ancestor of cultivated buckwheat, <it>Fagopyrum esculentum </it>ssp. <it>ancestrale</it>. The sequence was rapidly determined using a previously described approach that utilized a PCR-based method and employed universal primers, designed on the scaffold of multiple sequence alignment of chloroplast genomes. The gene content and order in buckwheat chloroplast genome is similar to <it>Spinacia oleracea</it>. However, some unique structural differences exist: the presence of an intron in the <it>rpl2 </it>gene, a frameshift mutation in the <it>rpl23 </it>gene and extension of the inverted repeat region to include the <it>ycf1 </it>gene. Phylogenetic analysis of 61 protein-coding gene sequences from 44 complete plastid genomes provided strong support for the sister relationships of Caryophyllales (including Polygonaceae) to asterids. Further, our analysis also provided support for <it>Amborella </it>as sister to all other angiosperms, but interestingly, in the bayesian phylogeny inference based on first two codon positions <it>Amborella </it>united with Nymphaeales.</p> <p>Conclusion</p> <p>Comparative genomics analyses revealed that the <it>Fagopyrum </it>chloroplast genome harbors the characteristic gene content and organization as has been described for several other chloroplast genomes. However, it has some unique structural features distinct from previously reported complete chloroplast genome sequences. Phylogenetic analysis of the dataset, including this new sequence from non-core Caryophyllales supports the sister relationship between Caryophyllales and asterids.</p

Comparative Analysis of Predicted Plastid-Targeted Proteomes of Sequenced Higher Plant Genomes

Plastids are actively involved in numerous plant processes critical to growth, development and adaptation. They play a primary role in photosynthesis, pigment and monoterpene synthesis, gravity sensing, starch and fatty acid synthesis, as well as oil, and protein storage. We applied two complementary methods to analyze the recently published apple genome (Malus x domestica) to identify putative plastid-targeted proteins, the first using TargetP and the second using a custom workflow utilizing a set of predictive programs. Apple shares roughly 40% of its 10,492 putative plastid-targeted proteins with that of the Arabidopsis (Arabidopsis thaliana) plastid-targeted proteome as identified by the Chloroplast 2010 project and ~57% of its entire proteome with Arabidopsis. This suggests that the plastid-targeted proteomes between apple and Arabidopsis are different, and interestingly alludes to the presence of differential targeting of homologs between the two species. Co-expression analysis of 2,224 genes encoding putative plastid-targeted apple proteins suggests that they play a role in plant developmental and intermediary metabolism. Further, an inter-specific comparison of Arabidopsis, Prunus persica (Peach), Malus x domestica (Apple), Populus trichocarpa (Black cottonwood), Fragaria vesca (Woodland Strawberry), Solanum lycopersicum (Tomato) and Vitis vinifera (Grapevine) also identified a large number of novel species-specific plastid-targeted proteins. This analysis also revealed the presence of alternatively targeted homologs across species. Two separate analyses revealed that a small subset of proteins, one representing 289 protein clusters and the other 737 unique protein sequences, are conserved between seven plastid-targeted angiosperm proteomes. Majority of the novel proteins were annotated to play roles in stress response, transport, catabolic processes, and cellular component organization. Our results suggest that the current state of knowledge regarding plastid biology, preferentially based on model systems is deficient. New plant genomes are expected to enable the identification of potentially new plastid-targeted proteins that will aid in studying novel roles of plastids

Genome editing in fruit, ornamental, and industrial crops

The advent of genome editing has opened new avenues for targeted trait enhancement in fruit, ornamental, industrial, and all specialty crops. In particular, CRISPR-based editing systems, derived from bacterial immune systems, have quickly become routinely used tools for research groups across the world seeking to edit plant genomes with a greater level of precision, higher efficiency, reduced off-target effects, and overall ease-of-use compared to ZFNs and TALENs. CRISPR systems have been applied successfully to a number of horticultural and industrial crops to enhance fruit ripening, increase stress tolerance, modify plant architecture, control the timing of flower development, and enhance the accumulation of desired metabolites, among other commercially-important traits. As editing technologies continue to advance, so too does the ability to generate improved crop varieties with non-transgenic modifications; in some crops, direct transgene-free edits have already been achieved, while in others, T-DNAs have successfully been segregated out through crossing. In addition to the potential to produce non-transgenic edited crops, and thereby circumvent regulatory impediments to the release of new, improved crop varieties, targeted gene editing can speed up trait improvement in crops with long juvenile phases, reducing inputs resulting in faster market introduction to the market. While many challenges remain regarding optimization of genome editing in ornamental, fruit, and industrial crops, the ongoing discovery of novel nucleases with niche specialties for engineering applications may form the basis for additional and potentially crop-specific editing strategies.The authors would like to acknowledge funding from MINECO, Spain (PGC2018-097655-B-I00 to P Christou), Generalitat de Catalunya Grant 2017 SGR 828 to the Agricultural Biotechnology and Bioeconomy Unit (ABBU). Work in the Dhingra lab in crop improvement is supported in part by Washington State University Agriculture Research Center Hatch grant WNP00011. ES and FR acknowledge the support received from the Department of Horticulture, BW was supported in part by a Research Assistantship from the Washington State University Graduate School. The authors would also like to thank Drs A. McHughen and H. Quemada for input and clarifications on US genome editing regulations. We would also like to thank the anonymous reviewers for their insightful comments

Transposons played a major role in the diversification between the closely related almond and peach genomes: Results from the almond genome sequence

We sequenced the genome of the highly heterozygous almond Prunus dulcis cv. Texas combining short and long‐read sequencing. We obtained a genome assembly totaling 227.6 Mb of the estimated 238 Mb almond genome size, of which 91% is anchored to eight pseudomolecules corresponding to its haploid chromosome complement, and annotated 27,969 protein‐coding genes and 6,747 non‐coding transcripts. By phylogenomic comparison with the genomes of 16 additional close and distant species we estimated that almond and peach (P. persica) diverged around 5.88 Mya. These two genomes are highly syntenic and show a high degree of sequence conservation (20 nucleotide substitutions/kb). However, they also exhibit a high number of presence/absence variants, many attributable to the movement of transposable elements (TEs). TEs have generated an important number of presence/absence variants between almond and peach, and we show that the recent history of TE movement seems markedly different between them. TEs may also be at the origin of important phenotypic differences between both species, and in particular, for the sweet kernel phenotype, a key agronomic and domestication character for almond. Here we show that in sweet almond cultivars, highly methylated TE insertions surround a gene involved in the biosynthesis of amygdalin, whose reduced expression has been correlated with the sweet almond phenotype. Altogether, our results suggest a key role of TEs in the recent history and diversification of almond and its close relative peach.info:eu-repo/semantics/publishedVersio