CRISPR/Cas9-mediated mutagenesis of WRKY3 and WRKY4 function decreases salt and Me-JA stress tolerance in Arabidopsis thaliana.
Arabidopsis
/ genetics
Arabidopsis Proteins
/ genetics
CRISPR-Cas Systems
Droughts
Gene Expression
/ genetics
Gene Expression Regulation, Plant
/ genetics
Genetic Engineering
/ methods
Plant Proteins
/ genetics
Plants, Genetically Modified
/ genetics
Polymorphism, Single Nucleotide
/ genetics
Salt Stress
/ genetics
Salt Tolerance
/ genetics
Sodium Chloride
/ metabolism
Stress, Physiological
/ genetics
Transcription Factors
/ genetics
Arabidopsis thaliana
CRISPR/Cas9 system
Me-JA stress
Salt stress
WRKY transcription factors
Journal
Molecular biology reports
ISSN: 1573-4978
Titre abrégé: Mol Biol Rep
Pays: Netherlands
ID NLM: 0403234
Informations de publication
Date de publication:
Aug 2021
Aug 2021
Historique:
received:
20
02
2021
accepted:
02
07
2021
pubmed:
6
8
2021
medline:
27
1
2022
entrez:
5
8
2021
Statut:
ppublish
Résumé
WRKY transcription factor is involved in regulation of plant growth and development, response to biotic and abiotic stresses, including homologous WRKY3 and WRKY4 genes which play a vital role in regulating plants defense response to pathogen and drought stress. To investigate the function of AtWRKY3 and AtWRKY4 genes in regulating salt and Me-JA stresses, the loss-of-function mutations were generated by clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated 9 (Cas9) system in Arabidopsis thaliana. Several independent transgenic lines with single or double mutations were obtained via Agrobacterium-mediated transformation. The knockout lines of AtWRKY3 and AtWRKY4 genes were successfully achieved and confirmed by qRT-PCR technology. Expression analysis showed that AtWRKY3 and AtWRKY4 genes had significantly up-regulated under salt and Me-JA stresses. The growth of double mutant plants under salt or Me-JA stresses were significantly inhibited compared with corresponding wild type (WT) plants, especially their root lengths. Moreover, the double mutant plants displayed salt and Me-JA sensitivity phenotypic characteristics, such as the increased relative electrolyte leakage (REL) and a substantial reduce in the activities of antioxidant enzymes, including superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) activities. Taken together, these data suggested that the simultaneous modification of homologous gene copies of WRKY are established using CRISPR/Cas9 system in A. thaliana and the loss of AtWRKY3 and AtWRKY4 has an effect on ROS scavenging pathways to reduce stress tolerance.
Sections du résumé
BACKGROUND
BACKGROUND
WRKY transcription factor is involved in regulation of plant growth and development, response to biotic and abiotic stresses, including homologous WRKY3 and WRKY4 genes which play a vital role in regulating plants defense response to pathogen and drought stress.
METHODS AND RESULTS
RESULTS
To investigate the function of AtWRKY3 and AtWRKY4 genes in regulating salt and Me-JA stresses, the loss-of-function mutations were generated by clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated 9 (Cas9) system in Arabidopsis thaliana. Several independent transgenic lines with single or double mutations were obtained via Agrobacterium-mediated transformation. The knockout lines of AtWRKY3 and AtWRKY4 genes were successfully achieved and confirmed by qRT-PCR technology. Expression analysis showed that AtWRKY3 and AtWRKY4 genes had significantly up-regulated under salt and Me-JA stresses. The growth of double mutant plants under salt or Me-JA stresses were significantly inhibited compared with corresponding wild type (WT) plants, especially their root lengths. Moreover, the double mutant plants displayed salt and Me-JA sensitivity phenotypic characteristics, such as the increased relative electrolyte leakage (REL) and a substantial reduce in the activities of antioxidant enzymes, including superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) activities.
CONCLUSION
CONCLUSIONS
Taken together, these data suggested that the simultaneous modification of homologous gene copies of WRKY are established using CRISPR/Cas9 system in A. thaliana and the loss of AtWRKY3 and AtWRKY4 has an effect on ROS scavenging pathways to reduce stress tolerance.
Identifiants
pubmed: 34351541
doi: 10.1007/s11033-021-06541-4
pii: 10.1007/s11033-021-06541-4
doi:
Substances chimiques
Arabidopsis Proteins
0
Plant Proteins
0
Transcription Factors
0
WRKY3 protein, Arabidopsis
0
WRKY4 protein, Arabidopsis
0
Sodium Chloride
451W47IQ8X
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
5821-5832Subventions
Organisme : Shanghai Sailing Program
ID : 19YF1414800
Informations de copyright
© 2021. The Author(s), under exclusive licence to Springer Nature B.V.
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