Mutation of GmAITR Genes by CRISPR/Cas9 Genome Editing Results in Enhanced Salinity Stress Tolerance in Soybean

Breeding of stress-tolerant plants is able to improve crop yield under stress conditions, whereas CRISPR/Cas9 genome editing has been shown to be an efficient way for molecular breeding to improve agronomic traits including stress tolerance in crops. However, genes can be targeted for genome editing to enhance crop abiotic stress tolerance remained largely unidentified. We have previously identified abscisic acid (ABA)-induced transcription repressors (AITRs) as a novel family of transcription factors that are involved in the regulation of ABA signaling, and we found that knockout of the entire family of AITR genes in Arabidopsis enhanced drought and salinity tolerance without fitness costs. Considering that AITRs are conserved in angiosperms, AITRs in crops may be targeted for genome editing to improve abiotic stress tolerance. We report here that mutation of GmAITR genes by CRISPR/Cas9 genome editing leads to enhanced salinity tolerance in soybean. By using quantitative RT-PCR analysis, we found that the expression levels of GmAITRs were increased in response to ABA and salt treatments. Transfection assays in soybean protoplasts show that GmAITRs are nucleus proteins, and have transcriptional repression activities. By using CRISPR/Cas9 to target the six GmAITRs simultaneously, we successfully generated Cas9-free gmaitr36 double and gmaitr23456 quintuple mutants. We found that ABA sensitivity in these mutants was increased. Consistent with this, ABA responses of some ABA signaling key regulator genes in the gmaitr mutants were altered. In both seed germination and seedling growth assays, the gmaitr mutants showed enhanced salt tolerance. Most importantly, enhanced salinity tolerance in the mutant plants was also observed in the field experiments. These results suggest that mutation of GmAITR genes by CRISPR/Cas9 is an efficient way to improve salinity tolerance in soybean.

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Genome Editing to Integrate Seed Size and Abiotic Stress Tolerance Traits in Arabidopsis Reveals a Role for DPA4 and SOD7 in the Regulation of Inflorescence Architecture

International Journal of Molecular Sciences ◽

10.3390/ijms20112695 ◽

2019 ◽

Vol 20 (11) ◽

pp. 2695 ◽

Cited By ~ 5

Author(s):

Siyu Chen ◽

Na Zhang ◽

Qimeng Zhang ◽

Ganghua Zhou ◽

Hainan Tian ◽

...

Keyword(s):

Abiotic Stress ◽

Stress Tolerance ◽

Genome Editing ◽

Seed Size ◽

Agronomic Traits ◽

Abiotic Stress Tolerance ◽

Loss Of Function ◽

Inflorescence Architecture ◽

Triple Mutant ◽

Transcription Repressors

Both seed size and abiotic stress tolerance are important agronomic traits in crops. In Arabidopsis, two closely related transcription repressors DPA4 (Development-Related PcG Target in the APEX4)/NGAL3 and SOD7 (Suppressor of da1-1)/NGAL2 (NGATHA-like protein) function redundantly to regulate seed size, which was increased in the dpa4 sod7 double mutants. Whereas ABA-induced transcription repressors (AITRs) are involved in the regulation of ABA signaling and abiotic stress tolerance, Arabidopsis aitr2 aitr5 aitr6 (aitr256) triple mutant showed enhanced tolerance to drought and salt. Here we performed CRISPR/Cas9 genome editing to disrupt DPA4 and SOD7 in aitr256 mutant, trying to integrate seed size and abiotic stress tolerance traits in Arabidopsis, and also to examine whether DPA4 and SOD7 may regulate other aspects of plant growth and development. Indeed, seed size was increased in the dpa4 sod7 aitr256 quintuple mutants, and enhanced tolerance to drought was observed in the mutants. In addition, we found that shoot branching was affected in the dpa4 sod7 aitr256 mutants. The mutant plants failed to produce secondary branches, and flowers/siliques were distributed irregularly on the main stems of the plants. Floral organ number and fertility were also affected in the dpa4 sod7 aitr256 mutant plants. To examine if these phenotypes were dependent on loss-of-function of AITRs, dpa4 sod7 double mutants were generated in Col wild type background, and we found that the dpa4 sod7 mutant plants showed a phenotype similar to the dpa4 sod7 aitr256 quintuple mutants. Taken together, our results indicate that the integration of seed size and abiotic stress tolerance traits by CRISPR/Cas9 editing was successful, and our results also revealed a role of DPA4 and SOD7 in the regulation of inflorescence architecture in Arabidopsis.

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Knockout of the entire family of AITR genes in Arabidopsis leads to enhanced drought and salinity tolerance without fitness costs

BMC Plant Biology ◽

10.1186/s12870-021-02907-9 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Siyu Chen ◽

Na Zhang ◽

Ganghua Zhou ◽

Saddam Hussain ◽

Sajjad Ahmed ◽

...

Keyword(s):

Abiotic Stress ◽

Stress Tolerance ◽

Abiotic Stresses ◽

Molecular Breeding ◽

Agronomic Traits ◽

Abiotic Stress Tolerance ◽

Plant Responses ◽

Fitness Costs ◽

Entire Family ◽

Knock Out

Abstract Backgorund Environmental stresses including abiotic stresses and biotic stresses limit yield of plants. Stress-tolerant breeding is an efficient way to improve plant yield under stress conditions. Genome editing by CRISPR/Cas9 can be used in molecular breeding to improve agronomic traits in crops, but in most cases, with fitness costs. The plant hormone ABA regulates plant responses to abiotic stresses via signaling transduction. We previously identified AITRs as a family of novel transcription factors that play a role in regulating plant responses to ABA and abiotic stresses. We found that abiotic stress tolerance was increased in the single, double and triple aitr mutants. However, it is unclear if the increased abiotic stress tolerance in the mutants may have fitness costs. Results We report here the characterization of AITRs as suitable candidate genes for CRISPR/Cas9 editing to improve plant stress tolerance. By using CRISPR/Cas9 to target AITR3 and AITR4 simultaneously in the aitr256 triple and aitr1256 quadruple mutants respectively, we generated Cas9-free aitr23456 quintuple and aitr123456 sextuple mutants. We found that reduced sensitivities to ABA and enhanced tolerance to drought and salt were observed in these mutants. Most importantly, plant growth and development was not affected even in the aitr123456 sextuple mutants, in whom the entire AITR family genes have been knocked out, and the aitr123456 sextuple mutants also showed a wild type response to the pathogen infection. Conclusions Our results suggest that knockout of the AITR family genes in Arabidopsis enhanced abiotic stress tolerance without fitness costs. Considering that knock-out a few AITRs will lead to enhanced abiotic stress tolerance, that AITRs are widely distributed in angiosperms with multiple encoding genes, AITRs may be targeted for molecular breeding to improve abiotic stress tolerance in plants including crops.

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CRISPR-mediated genome editing in maize for improved abiotic stress tolerance.

Molecular breeding in wheat, maize and sorghum: strategies for improving abiotic stress tolerance and yield ◽

10.1079/9781789245431.0023 ◽

2021 ◽

pp. 405-420

Author(s):

Ali Razzaq ◽

Ghulam Mustafa ◽

Muhammad Amjad Ali ◽

Muhammad Sarwar Khan ◽

Faiz Ahmad Joyia

Keyword(s):

Abiotic Stress ◽

Stress Tolerance ◽

Genome Editing ◽

Abiotic Stresses ◽

Abiotic Stress Tolerance ◽

Maize Yield ◽

Maize Genome ◽

Yield Improvement ◽

Maize Cultivars ◽

Genome Manipulation

Abstract This chapter discusses the applications of CRISPR-mediated genome editing to improve the abiotic stress tolerance (such as drought, heat, waterlogging and cold tolerance) of maize. CRISPR/Cas9 has great potential for maize genome manipulation at desired sites. By using CRISPR/Cas9-mediated genome editing, numerous genes can be targeted to produce elite maize cultivars that minimize the challenges of abiotic stresses. In the future, more precise and accurate variants of the CRISPR/Cas9 toolbox are expected to be used for maize yield improvement.

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Physiological and Molecular Characterization of Crop Resistance to Abiotic Stresses

Agronomy ◽

10.3390/agronomy10091308 ◽

2020 ◽

Vol 10 (9) ◽

pp. 1308

Author(s):

Monica Boscaiu ◽

Ana Fita

Keyword(s):

Abiotic Stress ◽

Stress Tolerance ◽

Crop Yields ◽

Agronomic Traits ◽

Abiotic Stress Tolerance ◽

Genetic Material ◽

Morphological Characteristics ◽

Stress Factors ◽

Special Issue ◽

Heat And Cold Stress

Abiotic stress represents a main constraint for agriculture, affecting plant growth and productivity. Drought and soil salinity, especially, are major causes of reduction of crop yields and food production worldwide. It is not unexpected, therefore, that the study of plant responses to abiotic stress and stress tolerance mechanisms is one of the most active research fields in plant biology. This Special Issue compiles 22 research papers and 4 reviews covering different aspects of these responses and mechanisms, addressing environmental stress factors such as drought, salinity, flooding, heat and cold stress, deficiency or toxicity of compounds in the soil (e.g., macro and micronutrients), and combination of different stresses. The approaches used are also diverse, including, among others, the analysis of agronomic traits based on morphological characteristics, physiological and biochemical studies, and transcriptomics or transgenics. Despite its complexity, we believe that this Special Issue provides a useful overview of the topic, including basic information on the mechanisms of abiotic stress tolerance as well as practical aspects such as the alleviation of the deleterious effects of stress by different means, or the use of local landraces as a source of genetic material adapted to combined stresses. This knowledge should help to develop the agriculture of the (near) future, sustainable and better adapted to the conditions ahead, in a scenario of global warming and environmental pollution.

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Kckout of the Entire Family of AITR Genes in Arabidopsis Leads to Enhanced Abiotic Stress Tolerance Without Fitness Costs

10.46678/pb.20.1055807 ◽

2020 ◽

Author(s):

Siyu Chen

Keyword(s):

Abiotic Stress ◽

Stress Tolerance ◽

Abiotic Stress Tolerance ◽

Fitness Costs ◽

Entire Family

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Molecular Abiotic Stress Tolerans Strategies: From Genetic Engineering to Genome Editing Era

10.5772/intechopen.94505 ◽

2020 ◽

Author(s):

Sinan Meriç ◽

Alp Ayan ◽

Çimen Atak

Keyword(s):

Abiotic Stress ◽

Genetic Engineering ◽

Stress Tolerance ◽

Genome Editing ◽

Target Genes ◽

Abiotic Stress Tolerance ◽

Industrial Applications ◽

Genomic Engineering ◽

Transporter Protein ◽

Abiotic Stress Factors

In last decades, plants were increasingly subjected to multiple environmental abiotic stress factors as never before due to their stationary nature. Excess urbanization following the intense industrial applications introduced combinations of abiotic stresses as heat, drought, salinity, heavy metals etc. to plants in various intensities. Technological advancements brought novel biotechnological tools to the abiotic stress tolerance area as an alternative to time and money consuming traditional crop breeding activities as well as they brought vast majority of the problem themselves. Discoveries of single gene (as osmoprotectant, detoxyfying enzyme, transporter protein genes etc.) and multi gene (biomolecule synthesis, heat shock protein, regulatory transcription factor and signal transduction genes etc.) targets through functional genomic approaches identified abiotic stress responsive genes through EST based cDNA micro and macro arrays. In nowadays, genetic engineering and genome editing tools are present to transfer genes among different species and modify these target genes in site specific, even single nuclotide specific manner. This present chapter will evaluate genomic engineering approaches and applications targeting these abiotic stress tolerance responsive mechanisms as well as future prospects of genome editing applications in this field.

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Genomics Applied to the Analysis of Flowering Time, Abiotic Stress Tolerance and Disease Resistance: A Review of What We Have Learned in Lolium spp.

Agriculture ◽

10.3390/agriculture10100425 ◽

2020 ◽

Vol 10 (10) ◽

pp. 425

Author(s):

Elisa Pasquali ◽

Gianni Barcaccia

Keyword(s):

Abiotic Stress ◽

Disease Resistance ◽

Stress Tolerance ◽

Flowering Time ◽

Magnaporthe Grisea ◽

Agronomic Traits ◽

Essential Feature ◽

Abiotic Stress Tolerance ◽

Gray Leaf Spot ◽

Forage Crops

Flowering time, abiotic stress tolerance and disease resistance are important agronomic traits of forage species like Lolium spp. Understanding the genetic control of these traits is enabled by the combination of genomic tools with conventional breeding techniques. Flowering time in this genus represents a complex trait due to the differences in the primary induction requirements among the species. In total, 36 QTLs (Quantitative Trait Locus) were identified across all seven linkage groups of Italian and perennial ryegrass involved in the flowering pathways, with several putative orthologous/homologous genes that have been characterized in other major crops. From the perspective of climate change, abiotic stress tolerance has become an essential feature; many QTLs that are involved in the control of plant responses have been identified, and transcriptional studies focusing on drought tolerance reported several DEGs (Differentially Expressed Genes) involved in carbon and lipid metabolism and signal transduction. Due to the incidence of microbial diseases, QTLs useful to developing cultivars resistant to bacterial wilt (Xanthomonas translucens pv. graminis), ryegrass crown rust (Puccinia coronata f. sp. Lolii) and gray leaf spot (Magnaporthe grisea/oryzae) have been mapped in both L. perenne and L. multiflorum populations. Due to the great importance of Lolium species, especially as forage crops, additional information about the three aforementioned agronomic traits is needed.

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