Genome-Wide Identification and Expression Profiling of Cytokinin Oxidase/Dehydrogenase (CKX) Genes Reveal Likely Roles in Pod Development and Stress Responses in Oilseed Rape (Brassica napus L.)

Abstract

Cytokinin oxidase/dehydrogenases (CKXs) play a critical role in the irreversible degradation of cytokinins, thereby regulating plant growth and development. Brassica napus is one of the most widely cultivated oilseed crops worldwide. With the completion of whole-genome sequencing of B. napus, genome-wide identification and expression analysis of the BnCKX gene family has become technically feasible. In this study, we identified 23 BnCKX genes and analyzed their phylogenetic relationships, gene structures, conserved motifs, protein subcellular localizations, and other properties. We also analyzed the expression of the 23 BnCKX genes in the B. napus cultivar Zhong Shuang 11 (‘ZS11’) by quantitative reverse-transcription polymerase chain reaction (qRT-PCR), revealing their diverse expression patterns. We selected four BnCKX genes based on the results of RNA-sequencing and qRT-PCR and compared their expression in cultivated varieties with extremely long versus short siliques. The expression levels of BnCKX5-1, 5-2, 6-1, and 7-1 significantly differed between the two lines and changed during pod development, suggesting they might play roles in determining silique length and in pod development. Finally, we investigated the effects of treatment with the synthetic cytokinin 6-benzylaminopurine (6-BA) and the auxin indole-3-acetic acid (IAA) on the expression of the four selected BnCKX genes. Our results suggest that regulating BnCKX expression is a promising way to enhance the harvest index and stress resistance in plants.

Role and Regulation of Cytokinins in Plant Response to Drought Stress

“…The other important component of CK metabolism is the CKX enzyme, which accounts for the irreversible inactivation of CKs by cleaving the side chain from CK molecules [46]. Genome-wide association studies have recently shed more light on the potential function of CKX members in plant species, with 13 CKX GFMs in maize [71], 11 CKX GFMs in rice [66], 12 CKX GFMs in apple [67], CKX GFMs in wheat (Triticum aestivum) [72] and 23 CKX GFMs in rapeseed (Brassica napus) [73]. Increasing lines of evidence suggest that CKX GFMs play important roles in various plant physiological and developmental modifications under drought stress [74][75][76] (Table 1).…”

Root enhancement in cytokinin-deficient oilseed rape causes leaf mineral enrichment, increases the chlorophyll concentration under nutrient limitation and enhances the phytoremediation capacity

“…In the genome of B. napus a large number of cytokinin metabolism and signalling genes was identified. Among others, there are 26 cytokinin-synthesizing IPT and 23 CKX genes , which are differentially expressed [37, 38] as compared to nine IPT and seven CKX genes in Arabidopsis. The fact that a large number of cytokinin genes have been retained in the Brassica genome suggests that the hormone exerts numerous distinct functions in this species.…”

Targeting Cytokinin Homeostasis in Rapid Cycling Brassica rapa with Plant Growth Regulators INCYDE and TD-K

“…Previous studies have predicted the location of Brassica rapa ssp. Pekinensis BrCKX3 protein in chloroplasts and secretory pathways [ 74 ], while in Brassica napus L. BnCKX3 was predicted to be in the vacuole [ 77 ], and confirmed to be vacuole-based in Arabidopsis [ 4 ]. However, the over-production of vacuole-located AtCKXs (AtCKX1 and AtCKX3) is known to cause severe traits associated with cytokinin deficiency in Arabidopsis [ 4 ], highlighting the potential importance of BrCKX3 in regulating cytokinin homeostasis and cytokinin sink dynamics at an intracellular level.…”