Loss‐function mutants of <i>OsCKX</i> gene family based on CRISPR‐Cas systems revealed their diversified roles in rice

Zheng, Xuelian; Zhang, Shuting; Liang, Yanling; Zhang, Rui; Liu, Li; Qin, Pengchen; Zhang, Zhe; Wang, Yan; Zhou, Jianping; Tang, Xu; Zhang, Yong

doi:10.1002/tpg2.20283

Cited by 15 publications

(11 citation statements)

References 93 publications

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“…The CKX gene family was identified or deduced in different plants, such as Arabidopsis, maize, cotton, and alfalfa. There are 7 AtCKX genes in Arabidopsis, 13 ZmCKXs in maize, 11 OsCKXs in rice, 8 VvCKXs in grape (Vitis vinifera L.), and 5 MnCKXs in mulberry (Morus notabilis) [7,[26][27][28][29]. In this study, five LcCKX genes were identified in L. chinensis.…”

Section: Discussionmentioning

confidence: 88%

Genome-Wide Identification and Abiotic-Stress-Responsive Expression of CKX Gene Family in Liriodendron chinense

Sun

Zhu

Hao

et al. 2023

Plants

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Liriodendron chinense is a tree species of the Magnoliaceae family, an ancient relict plant mainly used for landscaping and timber production due to its excellent material properties and ornamental value. The cytokinin oxidase/dehydrogenase (CKX) enzyme regulates cytokinin levels and plays an important role in plant growth, development, and resistance. However, too-high or too-low temperatures or soil drought can limit the growth of L. chinense, representing a key issue for research. Here, we identified the CKX gene family in the L. chinense genome and examined its transcriptional responses to cold, drought, and heat stresses. A total of five LcCKX genes, distributed on four chromosomes and divided into three phylogenetic groups, were identified across the whole L. chinense genome. Further analysis showed that multiple hormone- and stress-responsive cis-acting elements are located in the promoter regions of LcCKXs, indicating a potential role of these LcCKXs in plant growth, development, and response to environmental stresses. Based on existing transcriptome data, LcCKXs, especially LcCKX5, were found to transcriptionally respond to cold, heat, and drought stresses. Furthermore, quantitative reverse-transcription PCR (qRT-PCR) showed that LcCKX5 responds to drought stress in an ABA-dependent manner in stems and leaves and in an ABA-independent manner in roots. These results lay a foundation for functional research on LcCKX genes in the resistance breeding of the rare and endangered tree species of L. chinense.

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Section: Discussionmentioning

confidence: 88%

Genome-Wide Identification and Abiotic-Stress-Responsive Expression of CKX Gene Family in Liriodendron chinense

Sun

Zhu

Hao

et al. 2023

Plants

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“…The composition of the entrance triplet likely co-relates with other sequential (and consequently structural) features responsible for additional fine-tuning of the protein’s substrate specificity. The ability of CKXs to discriminate CKs with different N6 and N9 substitutions could also contribute to the varying effects of OsCKX mutations on endogenous concentrations of tZR, iP, and iPR in rice (Zheng et al ., 2023). Our results suggest that the diversity of CKX isoforms and their substrate specificities is conserved among the monocot species, if not the plant kingdom as a whole, considering that similar trends were observed in Arabidopsis (Frébortová et al ., 2007; Galuszka et al ., 2007; Kowalska et al ., 2010).…”

Section: Discussionmentioning

confidence: 99%

Cytokinin Dehydrogenase in Xylem Sap Reveals A Direct Link Between Cytokinin Metabolism and Long-Distance Transport

Nedvěd,

Motyka,

Raimanová

et al. 2023

Preprint

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Metabolic degradation of plant hormones cytokinins (CKs) co-regulates their homeostasis and signalling. In this work, we employed a large-scale bioinformatical analysis to address a diversity of cytokinin oxidase/dehydrogenase (CKX) substrate specificities previously described in several case studies. We present a three-way correlation of the entire CKX amino acid sequences, a variable motif involved in substrate binding, and subcellular localizations predicted by a deep learning model. This correlation is conserved in monocotyledonous plants, suggesting that the CKX diversity in a single species allows a precise tuning of the CK homeostasis. Following these findings, we detected CKX activity in xylem sap for the first time, using the oat (Avena sativa) as a model plant. Further investigation of the substrate specificity and glycosylation of this xylem-located CKX suggested that it originates in roots. We have identified 27 putative CKXs in oats and attributed the xylem-located activity to the extracellular isoforms AsCKX1a,c,d. Finally, we show that the xylem-located CKX activity responds to the nitrate supply, highlighting its physiological relevance. Taken together, we show that CKX directly modulates root-to-shoot CK translocation through metabolic degradation of the transported CKs.

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“…Agrobacterium-mediated rice transformation was conducted by following an modified protocol 66,67 . In brief, calli derived from rice seeds were induced in a growth chamber for 7 days at 32°C under light.…”

Section: Methodsmentioning

confidence: 99%

High performance TadA-8e derived cytosine and dual base editors with undetectable off-target effects in plants

Fan,

Cheng,

et al. 2024

Preprint

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Cytosine base editors (CBEs) and adenine base editors (ABEs) are powerful tools to install precise C-to-T and A-to-G base edits in living organisms. In recently years, a highly efficient ABE based on engineered TadA-8e, known as ABE8e, has been developed to confer highly efficient A-to-G editing in mammalian cells and plants. Interestingly, TadA-8e can be further evolved and engineered to confer cytosine base editing. In this study, we compare in rice and tomato cells a serial of engineered CBEs derived from TadA-8e and identify TadCBEa, TadCBEd, and TadCBEd_V106W as efficient CBEs with high editing purity, while maintaining the relatively narrow editing window of ABE8e. Excitingly, we show TadDE is a new class of dual base editor where a single deaminase domain promotes simultaneous C-to-T and A-to-G base editing in plant cells. Multiplexed base editing is demonstrated in transgenic rice plants with TadCBEa and TadDE. Furthermore, no off-target effects of TadCBEa and TadDE can be detected by whole genome and transcriptome sequencing of the transgenic lines, suggesting high editing specificity. Finally, we demonstrate two gain-of-function crop engineering applications in rice by using the efficient dual base editor, TadDE. In the first case, herbicide resistance alleles ofOsALSare introduced by TadDE. In the second case, synonymous mutations are generated inOsSPL14, making its transcript resistant toOsMIR156-mediated degradation. Together, this study reports TadA-8e derived CBEs and a dual base editor with high editing activity, purity, and specificity. They are valuable additions to the expanding plant base editing toolbox.

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Loss‐function mutants of OsCKX gene family based on CRISPR‐Cas systems revealed their diversified roles in rice

Cited by 15 publications

References 93 publications

Genome-Wide Identification and Abiotic-Stress-Responsive Expression of CKX Gene Family in Liriodendron chinense

Genome-Wide Identification and Abiotic-Stress-Responsive Expression of CKX Gene Family in Liriodendron chinense

Cytokinin Dehydrogenase in Xylem Sap Reveals A Direct Link Between Cytokinin Metabolism and Long-Distance Transport

High performance TadA-8e derived cytosine and dual base editors with undetectable off-target effects in plants

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