CARBON Newsletter (3 May) - Your Latest News About CRISPR in AgroBio

Top picks

• Federal authorities in Belgium have granted permission to perform three new field trials with CRISPR gene-edited maise that researchers at VIB have developed. The plants in question have 1) improved tolerance to DNA damage caused by UV-light thanks to a mutation in the transcription factor ZmNAC52, 2) better digestibility through reduced content of lignin in the cell walls, and 3) improved tolerance to prolonged drought through increased chromatin condensation that causes growth to pause.
• Enhanced rice grain yield has been obtained by overcoming the trait tradeoff between panicle number and size. The achievement was made through CRISPR-based targeted deletion of a 54-base pair cis-regulatory element in the Ideal Plant Architecture 1 (IPA1) gene that increases grains per panicle but reduces tillers. The Chinese researchers suggest that targeting gene regulatory regions should help dissect tradeoff effects and provide a rich source of targets for breeding complementary beneficial traits.

Technical advances

• A human study with potential applications for agriculture demonstrated targeted A-to-G base editing in mitochondrial DNA with a new class of programmable deaminases called transcription-activator-like effector (TALE)-linked deaminases (TALEDs). Custom-designed TALEDs were highly efficient in human cells, catalysing A-to-G conversions at 17 target sites in various mitochondrial genes with editing frequencies of up to 49%. The researchers from South Korea are also focusing on developing TALEDs suitable for A-to-G base editing in chloroplast DNA, which encodes essential genes in photosynthesis in plants.
• Chinese researchers have identified seven U6 RNAs from eggplant (Solanum melongena) and cloned their promoters. Either these SmU6 promoters or the typical AtU6 promoters from Arabidopsis were used to drive the expression of gRNAs in CRISPR-Cas9 vectors for gene editing of eggplant cotyledons. Results indicated that the editing efficiency of eggplant U6 promoters was better than that of Arabidopsis U6 promoters.

Disease control

• Researchers in Kazakhstan have obtained virus resistance in five local elite barley cultivars by CRISPR-Cas9 gene editing of the eukaryotic translation initiation factor 4E (eIF4E). Cas9 and gRNA were delivered into barley cells via a proprietary Agrobacterium germ-line transformation technique.
• A not-yet peer-reviewed paper on the bioRxiv server describes the use of CRISPR-Cas9 to develop bacterial blight-resistant cassava plants. The Chinese researchers targeted the MeSWEET10a promoter to decrease sucrose transport from the interior of plant cells to the apoplasts and thereby exclude the bacteria from its carbon sources. As a result, stable and robust resistance was obtained with no significant differences in tuber root traits and starch content.

Nutritional quality

• Researchers in the UK demonstrate the utility of dual-guided CRISPR-Cas9 to control the expression of a gene by deleting an intergenic sequence. The approach was used to fuse the promoter of a non-essential photosynthesis-related gene to the coding sequence of DIACYLGLYCEROL ACYLTRANSFERASE 2 (DGAT2) in the lipase-deficient sugar-dependent 1 mutant of Arabidopsis thaliana. Gene editing led to a more than twenty-fold increase in DGAT2 expression, and the triacylglycerol content increased by around thirty-fold.

Agronomic traits

• American researchers have developed alfalfa (Medicago sativa) that hyper accumulates phosphate at levels 3-6-fold higher than wild-type after CRISPR-Cas9 editing of PHOSPHATE2 (PHO2). The plants might have practical applications reclaiming Pi from contaminated or enriched soil and can be used in conservation buffer strips to protect waterways from Pi run-off.
• Researchers in China have improved salt stress resistance in rice by CRISPR-Cas9-mediated knockout of the OsbHLH024 transcription factor. A single nucleotide deletion resulted in a significant increase in the shoot weight, the total chlorophyll content, and the chlorophyll fluorescence when mutants were grown under salt stress. Mutants also showed better control of oxidative stress, fewer Na⁺ but more K⁺, and a balanced level of Ca²⁺, Zn²⁺, and Mg²⁺ in the shoots and roots.

Regulation and opinion

• CRISPR won’t work in the real world for farmers, researchers and all of agriculture. That's the bold statement from Lotte Westerhoff at Hudson River Biotechnology, and she gives three reasons for the failure: 1. Transgenes are ineffective and cause regulation issues, 2. Regeneration is difficult, and 3. CRISPR protein can entail high licensing costs.
• A technology feature in Nature highlights the achievements of Chinese researcher Caixia Gao who has made tremendous progress in gene editing in plants. The feature focuses on her efforts in developing more efficient base and prime editors to generate crops with greater disease resistance, higher nutritional content or more fruit per plant. Caixia Gao recently engineered a prime editor with enhanced editing efficiency in plants.
• The European Commission has opened a public consultation about the legislation for plants produced by certain new genomic techniques. Citizens both in and outside the EU are invited along with stakeholders to fill out an online questionnaire. Here, they will be asked about their views on the functioning of the current GMO legislation for plants obtained by targeted mutagenesis and cisgenesis and their food and feed products, i.e. under the current GMO legislation, and on potential options for a new framework.
• If you have a hard time understanding the EU's policy on gene-edited plants, there is help in this breakdown from the global law firm, Taylor Wessing. The piece explains how the EU is trying to catch up with the state of development of new techniques for genome-edited plants in a new legal framework.
• A feature in Giant Freakin Robot looks at the possibilities of using CRISPR to fight climate change. Among the strategies that scientists are taking is to gene-edit plants to absorb carbon dioxide and accumulate it as suberin - basically cork - in the roots.
• In an Agricultural Science & Technology editorial, researchers from the USA and China discuss advances in genome editing for sustainable agriculture. The editorial is part of a special issue that includes several articles and reviews about gene editing in plants that appeared in the journal earlier this year.

Reviews

• Chinese researchers review the application of the CRISPR-Cas9 technology in horticultural crops for stress response enhancement, fruit quality improvement, and cultivation traits modification. They also describe the construction and optimisation of the CRISPR-Cas9 system, including sgRNA promoter, sgRNA design, Cas9 protein promoter, SpCas9 variants and orthologs, and vector delivery methods.
• A mini-review by researchers from India discusses specific studies that highlight the prospects and tradeoffs associated with the choice of promoters for genome editing. Particular emphasis is on the need for inductive exploration and discovery to further advance this area of research in crop plants.
• Another review by Indian researchers looks at cytokinins and their role in sustainably increasing crop production through CRISPR-Cas gene editing of cytokinin metabolism. The review also summarises the current developments of RNP-mediated DNA/transgene-free genomic editing of plants.
• Yet another review from India highlights the CRISPR-Cas system's possible application for improving the acquisition and redistribution of nutrients from the soil. The review discusses the effects of engineering key residues of nutrient transporters, transcriptional regulation of nutrient transport signals, engineering motifs in promoters and transcription factors.
• Advancements and opportunities for rice improvement using CRISPR-based genome editing are the subjects of a review by Chinese researchers. The review covers the mechanisms of CRISPR-Cas9, including newly emerged Cas variants, vector construction, model design and different bioinformatics tools for target site selection.
• In a review, researchers in India discuss recent advancements in CRISPR-Cas technology for accelerated crop improvement. The review emphasises emerging gene-editing techniques like base and prime editing, multiplex genome editing, chromosome engineering and dCas9 mediated epigenetic modification.
• A review by Indian researchers looks at the molecular mechanism of drought response in plants and the application of the CRISPR-Cas genome-editing system for improving drought tolerance in plants.

Meetings and webinars

• The 2022 In Vitro Biology Meeting takes place 4-7 June in San Diego, California. Among the sessions are "Advanced Applications of Site-Directed Nucleases" and "Frontiers in Gene Editing for Crop Improvement", including several distinguished speakers.
• The 23rd Plant Biology Symposium - RNA Biology will take place 18-20 May at Penn State in the USA. The focus is on plants, and topics covered include small RNAs, RNA structure, RNA processing, localisation, translation, epigenetics, and interconnections to the roles that RNA plays in plant growth, development, physiology, defence, and abiotic stress response.