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Article

Cytokinin Oxidase (CKX) Family Members in Potato (Solanum tuberosum): Genome-Wide Identification and Expression Patterns at Seedling Stage under Stress

by
Wei Zhang
*,
Shangwu Liu
,
Shaopeng Wang
,
Feifei Xu
,
Zhenyu Liu
and
Bei Jia
Heilongjiang Academy of Agricultural Sciences, Harbin 150006, China
*
Author to whom correspondence should be addressed.
Horticulturae 2024, 10(7), 737; https://doi.org/10.3390/horticulturae10070737
Submission received: 30 May 2024 / Revised: 2 July 2024 / Accepted: 11 July 2024 / Published: 12 July 2024
(This article belongs to the Special Issue Genome-Wide Identification and Expression Analysis for the Genetic Improvement of Horticultural Plants)
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Abstract

:
Cytokinin (CK) is an important hormone that regulates cell differentiation. The CK content in plants is regulated by cytokinin oxidase (CKX), an important enzyme that participates in hormone-regulated pathways. Additionally, CKXs comprise a large family of enzymes, but little information exists on the CKXs in potato (Solanum tuberosum). In this study, nine CKXs were identified in the potato genome and named StCKX01-09, according to their order on the linkage groups (LGs). They belong to six subfamilies, and the members within the respective subfamilies had similar motifs, a similar gene structure, and similar cis-acting elements. Additionally, the CKXs from four other species, including Arabidopsis, rice (Oryza sativa), soybean (Glycine max), and maize (Zea mays), were also divided into six subfamilies, while members within each subfamily had similar types of motifs. Moreover, the potato StCKXs were shown to influence plant hormones and stress-related factors. StCKXs were collinear, with one CKX in Arabidopsis and five CKXs in Glycine max. Quantitative real-time PCR (qRT-PCR) revealed tissue-specific expression patterns in the potato seedlings and changes in the expression levels in response to stress. Furthermore, the cytokinin content and CKX enzyme activity were shown to be regulated by StCKXs. This study provides detailed information that can help future endeavors in the molecular breeding of potato (Solanum tuberosum).

1. Introduction

Plant hormones are an important class of regulatory substances involved in plant growth and development [1]. Cytokinins (CKs) are a group of plant hormones that promote cell division and differentiation [2], and their concentration can influence cell division, sprout formation, and overall growth in plants [3]. CK is generally produced in the plant roots and promotes cytoplasmic division and differentiation, and the growth of various tissues synergistically alongside other plant hormones like auxin [4,5]. CK is also involved in certain physiological activities, such as the germination of seeds, elongation of roots, and development of leaves [6]. Notably, CK has been documented to be located adjacent to the anti-codon in tRNA and participated in the junction of tRNA and ribosomal mRNA complexes during protein synthesis [7]. CK is synthesized through the modification of the original purines found in tRNA, although the mechanism underlying CK’s function is not yet fully understood [8]. Exogenous CK promotes the synthesis of nitrate reductase, proteins, and nucleic acids [9]. The main physiological functions of CK include the induction of cell division, sprout formation, and bud growth [10]. In tobacco tissue cultures, CK also induces the callus cells in the pulp or stem segments that had stopped dividing to begin dividing again, and this phenomenon has been used as a bioassay to detect the presence of CK [11]. In addition, CK inhibits adventitious root formation and lateral root formation and delays leaf senescence [12]. Some CK-related genes have been identified in the potato (Solanum tuberosum) genome and constitute the CK regulatory system, in which CK’s effects on tuber initiation are realized via their action in stolons [13].
CK degradation is catalyzed by cytokinin oxidase/dehydrogenase (CKX), which is currently the only enzyme known to specifically catalyze the irreversible degradation of natural isopentene CKs such as isopentenyl adenine [14]. CKX was first discovered in tobacco cells and later identified in wheat and Arabidopsis [15]. The structure of CKX plays a key role in its activity and has always been a focus of research. The FAD-binding region of CKX is the active site responsible for its enzyme activity and has a GHS (Glu-His-Ser (GHS) residue pattern. CKX covalently binds to FAD through His [16].
The CKX gene family is widely involved in plant growth and development through participation in CK’s metabolism. AtCKX1 (a CKX member in Arabidopsis) has been reported to have a strong effect on the roots, where the overexpression of the enzyme promotes below-ground plant growth while inhibiting aboveground growth [17]. Correspondingly, the CK concentration increased in ckx3/ckx5 double mutants relative to wild-type plants and resulted in an increased number of flower primordia, pods, and yield [18]. The expression level of OsCKX2 in this mutant was observed to be low, leading to the large accumulation of CKs in the floral primordia and an increase in the number of reproductive organs and biomass [19]. Additionally, the aboveground biomass increased when the CKX enzyme activity was inhibited in barley (Hordeum vulgare), but the root weight decreased [20]. These studies show that the function of CKXs is relatively conserved among the different species. The CKX family members have also been reported to participate in the plant’s responses to external environmental changes [21]. It was found that the expression levels of most members of the AtCKX gene family in Arabidopsis increased under drought or salt stress. MdCKX7.2 expression in apple was also increased in response to drought stress [22,23]. Some CKX genes have been cloned to investigate their function, and they were shown to be involved in plants’ responses to abiotic stress [24]. Similarly to AtCKX1 in Arabidopsis, the ectopic expression of CKX enzymes in transgenic tobacco significantly enhanced the plants’ drought resistance phenotype [25], while AtCKX1/2/3/4 overexpression in Arabidopsis increased salt stress resistance to varying degrees [26]. CKXs also exhibited various expression patterns in response to changes in the expression of other hormones [27]. The expression level of the CKXs increased when the plants were treated with exogenous 6-BA and abscisic acid (ABA) [28]. These studies show that CKX genes play an important role in plant growth and development, and in responses to external environmental changes.
Moreover, CKX gene family members have been identified in various plant species, such as Arabidopsis, rice (Oryza sativa), maize (Zea mays) [29], tobacco (Nicotiana tabacum) [30], wheat (Triticum aestivum) [31], common bean (Phaseolus vulgaris) [27], and foxtail millet (Setaria italica) [32]. Some studies have shown that StCKXs could affect cytokinin metabolism and tuber dormancy progression in potato (Solanum tuberosum) [33]. However, there have been limited studies on the function of CKXs in potato under stress conditions. The global potato production is the third largest among non-cereal crops [34], and second only to rice and wheat. Due to climate change, potatoes are frequently affected by various biotic and abiotic sources of stress during cultivation [35]. Therefore, the identification and utilization of stress-resistance genes in potato and the development of new stress-resistant breeds hold promising potential. This study identified CKX members in the reference genome of potato, and a comprehensive analysis was conducted to understand their characteristics. The study also provides detailed and essential insights that will be useful for the molecular breeding of CKX members in potato (Solanum tuberosum).

2. Materials and Methods

2.1. Identification of CKX Members in Potato

The protein and nucleotide sequences of the potato CKX members were obtained from the Ensembl plants database, and SolTub_3.0 (4113) was used as the reference genome. The CKX protein domain with the hmm domain number PF09265 [27] was downloaded from the Pfam database [36]. These CKXs were screened using the ExPASy Proteomics Server [37], and their chromosomal location was mapped relative to the reference genome using TBtools [38].

2.2. Analysis of the Characteristics of CKX Genes in Potato

To analyze the evolutionary relationship among the CKX members, a maximum likelihood evolutionary tree of the CKX protein sequences was built using MEGA X with a JTT+G model and 1000 bootstrap replicates [29,39]. During motif analysis, the sequences were filtered to include only those with motif lengths of 10–50 amino acids and e-values of less than 1 × 10−20 [40]. The Gene Structure Display Server software V2.0 was used to analyze the gene structure of the CKX members, including the coding sequence and intron sequence [41]. PlantCARE (https://bioinformatics.psb.ugent.be/webtools/plantcare/html/, accessed on 29 May 2024) was used to predict the presence and functions of cis-acting elements within CKX gene promoters in potato [42]. Regarding the motif analysis, MCScanX was used to identify putative homologous chromosomal regions (i.e., collinearity) among the potato genomes [43], and Circos was used for visualizing the results [44].

2.3. Plant Materials and Conditions

This study used the “YouJin” potato variety, a locally cultivated variety grown in Heilongjiang province, China. The plants were grown in a solar greenhouse in Harbin, China. The following treatments were set up at the seedling stage: salt stress (S) with 70 mmol/L NaCl, heavy metal stress (H) with 60 mg/L HgCl2, cold stress with 4 °C (C) for 48 h, and biological stress with Clavibacter michiganensis subsp. (Cms) [45]. The control plants were grown in regular soil., similar to the methods described in Zhang [46].

2.4. Measurement of the Cytokinin Content

The leaves in the stress treatments were collected after two days, while different tissues were used as samples in the tissue-specific analyses. The cytokinin content in each sample was determined using ELISA, and the CKX activity in each sample was measured using ELISA (Michy Biology, Nanjing, China).

2.5. The CKX Members’ Expression Levels

The CKX expression levels in the potato plants were measured using quantitative real-time PCR (qRT-PCR). More specifically, the total RNA in the respective samples was extracted using the TRIzol Plus RNA Purification Kit (Vendor, Covington, LA, USA). After assessing the quality of the purified RNA with the NanoDrop (OneC, Thermo, Emeryville, CA, USA) and electrophoresis, the RNA was used to synthesize single-stranded cDNA using the cDNA Synthesis Kit. The CKX-specific primers were designed using the Primer 5.0 software, in which St-actin was set as the reference gene (Table S1) [47]. The 2 × HotStart SYBR qPCR Master Mix was used to perform the qRT-PCR in triplicate in a LightCycler® thermocycler, with the following cycling conditions: 95 °C for 3 min and then 95 °C for 5 s, 60 °C for 30 s, and 72 °C for 20 s for 40 cycles. Then, the relative expression level of StCKXs was analyzed by the 2−ΔCt method according to the Ct values [48].

2.6. Data Analysis

SPSS version 19.0 was used to analyze the data generated in this study, which are presented as means ± standard deviation of three biological replicates. At the same time, the statistical differences (p < 0.05) were calculated between different treatment groups by two-way analysis of variance (ANOVA). The figures were drawn using Prism [49].

3. Results

3.1. Identification of CKX Family Members in the Potato Genome

The CKX genes were identified using hmm-search, and nine candidates remained after removing duplicates. These nine genes were designated StCKX01-StCKX09 according to their positions on the potato genome. The nine genes were located on five chromosomes, of which the chromosomes 4, 8, 10, and 12 harbored two and the chromosome 1 only one StCKX (Figure 1 and Table 1). The coding sequence (CDS) of the nine genes ranged from 276 to 2100 nucleotides in length, while the molecular weight ranged from 11,095.66 to 79,314.95. The theoretical pI of the enzymes was between 5.33 and 8.12, while the instability and aliphatic index results differed among the StCKXs.

3.2. Evolutionary Analysis of StCKXs

The evolutionary relationship among the StCKXs was investigated using MEGA X. The nine members were divided into six clusters, consisting of three pairs of StCKXs, and three other members who were relatively distant from the others (Figure 2).

3.3. Comparative Motif and Gene Structure Analysis of StCKXs

Ten motifs were found when the nine StCKX proteins were compared with the MEME Suite database. No motifs were found in StCKX06, while StCKX04 only had motifs 6 and 9. The other StCKX proteins all had motifs 1, 2, and 3, thereby suggesting that most StCKXs may have similar functions to proteins in the MEME database with the same predicted motifs (Figure 3A,B). When the gene structure was examined, the sequences of the StCKXs in cluster IV (StCKX08 and StCKX09) were longer than those in the other clusters. Interestingly, StCKX01, StCKX03, and StCKX04 had both the UTR and coding sequence (CDS), while the other StCKXs only had the CDS region. The results showed no motifs in StCKX06 (Figure 3C).

3.4. Comprehensive Analysis of the CKXs

The CKXs in Arabidopsis, soybean (Glycine max), maize (Zea mays), rice (Oryza sativa), and potato were identified on the reference genomes using the same Pfam hmm-search conditions. The CKXs of Arabidopsis, soybean, maize, rice, and potato were grouped into six major clusters, similar to the grouping of the potato StCKXs. The motif analysis of these CKXs showed that the members in the same evolutionary cluster had similar motif configurations. These results suggest that the CKXs from the five species that belong to the same clusters may have a similar structure and function (Figure 4).

3.5. Analysis of the cis-Acting Elements of StCKXs

In order to understand the function of StCKXs, the promoter regions of the StCKX genes were identified in the reference potato genome, with a 1500 bp sequence upstream of the CDS used as the promoter sequence. The 12 cis-acting elements identified in the promoter region of the genes could be divided into three functional types: hormone-related cis-acting elements, stress-related cis-acting elements, and sprout-related cis-acting elements. Six and five subtypes of hormone-related and stress-related elements were found, respectively. These results indicate that StCKXs may be involved in regulating hormone homeostasis and stress responses (Figure 5 and Table S2).

3.6. Collinearity Analysis of StCKXs

To understand the evolution of StCKXs and potentially predict their functions, the genetic relationship between the CKXs of potato and those of other species was investigated. Six pairs of collinear genes were found between the StCKXs and the CKXs from dicotyledonous plants, including Arabidopsis and soybean (Glycine max), of which five pairs were observed with soybean genes and one pair with an Arabidopsis gene. The difference in the number of collinear gene pairs may be due to the closer genetic relationship between the Solanaceae and Leguminosae families (Figure 6).

3.7. Gene Ontology Enrichment Analysis of StCKXs

The gene ontology database was examined to determine which GO terms were up-or down-regulated in relation to the StCKXs, using a q-value of less than 0.05 as a threshold. The top 20 results are shown in Table 2. Some of the GO terms, such as GO:0009690, GO:0010817, GO:0019139, and GO:0042445, were related to hormones, while the GO terms GO:0016614, GO:0055114, and GO:0016491 were related to oxidation. These results suggest that StCKXs may regulate plant growth by regulating hormone homeostasis and signaling and oxidation.

3.8. Tissue-Specific Expression Patterns of StCKXs

The expression patterns of the StCKXs in the roots, stems, and leaves of potato seedlings in tissue culture were investigated. The expression levels of most StCKXs, including StCKX01, StCKX02, StCKX03, StCKX05, and StCKX07, in the leaves were significantly higher than those in the roots and stems, while the expression levels of StCKX04 were similar in the leaves and stems. Only StCKX06 had a higher expression level in the stems than in the leaves and roots. The expression levels of StCKX08 and StCKX09 were higher in the leaves and roots than in the stems. These results indicate that the leaves can be used in further experiments under different conditions to elucidate the expression pattern differences among StCKXs (Figure 7).

3.9. Expression Patterns of StCKXs Exposed to Various Stress Condition

The expression levels of the StCKXs in the leaves were measured when they were submitted to various biotic and abiotic sources of stress. The results showed that the StCKX expression levels changed significantly (p < 0.05) when the plants were under stress. Some StCKXs, such as StCKX02 and StCKX03, had lower expression levels when under stress, while most others had higher expression levels (Figure 8). Taken together, these results showed that StCKXs respond to both biotic and abiotic stresses.
Moreover, the CKX activity and the CK content in the leaves under salt (S), heavy metal (H), cold (C), and Clavibacter michiganensis subsp. (Cms) stress were examined. The results showed that the CKX activity increased in accordance with the increase in the expression of certain StCKXs, as mentioned above, while the CK content decreased significantly (p < 0.05). These results showed that StCKXs may respond to stress-inducing conditions, resulting in changes in the cytokinin oxidase activity and cytokinin content, and these results are consistent with those observed in the GO enrichment analysis (Figure 9).

4. Discussion

CKX genes have been identified in various plant species, with diverse members identified through analyses of reference genomes [50]. A total of 7, 7, 10, and 13 CKX genes have been found in Arabidopsis, alfalfa, rice, and maize, respectively [29,41,51,52]. As many as 17 CKX genes have been found in soybean (Glycine max), which is considered to be more than in most other species [53]. Additionally, using different versions of a reference genome led to the identification of different numbers of CKX genes, such as in bread wheat, in which a range of 11–14 CKXs have been found in different versions of the reference genome [54]. The varying number of CKXs may be partly because the genome size often correlates with the number of discovered CKXs [55]. Moreover, genome evolution and gene duplication likely influence the number of CKXs harbored by the different plant species. In this study, nine CKXs were identified in the reference genome of potato and located on chromosomes 1, 4, 8, 10, and 12. The absence of CKXs on chromosomes 2 and 3 may be due to the low frequency of gene duplications among CKXs. Similarly, the CKXs in wheat (Triticum aestivum) and common bean (Phaseolus vulgaris) were not present on all the chromosomes of these genomes [27,50]. Regarding their evolutionary analysis, CKXs clustered into four to eight clusters [56], and the potato StCKXs were grouped within six of these clusters. It is worth noting that the JTT model also appeared to be particularly effective with CKXs in the evolutionary analysis. However, the variation in the subgroup numbers may be attributable to the number of CKXs in the respective genomes, which is similar to CKXs in soybean and common bean [57].
The motifs in the potato StCKXs belonging to the same subgroups were similar (Figure 3A,B), and a similar trend was observed among the CKXs in alfalfa and oilseed rape (Brassica napus) [58,59]. StCKXs promoters contained three types of cis-acting elements, and those within similar subgroups were also similar. Furthermore, certain hormone- and stress-related elements seemed to be features of CKX members [59,60], suggesting that StCKXs may be involved in the plant’s response to hormones and stress. The collinearity analysis revealed five StCKXs that were collinear with CKXs of soybean and one StCKX that was collinear with a gene from Arabidopsis. These observations suggest that CKXs from Solanaceae and Leguminosae crops may have similar gene and genome structures. Moreover, AT1G75450, which was collinear with StCKX03, has been reported to act upstream of the cytokinin catabolism pathway [61] and was responsive to infections by Botrytis cinerea. This suggests that StCKXs may respond to stress-related factors. CKXs showed tissue-specific variations in their expression levels in wheat and apple [45,51]. In potato, StCKXs also displayed specific expression patterns in different tissues at the seedling stage. The GO enrichment analysis of StCKXs showed that they may participate in cytokinin-related pathways, similar to the common bean [6]. Additionally, biotic and abiotic sources of stress have been shown to influence plants during all stages of growth and development. In contrast, the seedling stage has been reported to be the most sensitive to abiotic stress and directly affects development [57]. Cytokinin is an important class of plant hormone and is involved in the plant’s response to various biotic and abiotic sources of stress [60].
Moreover, the expression patterns of CKX members revealed their tissue specificity and responsiveness to stimuli [56]. Understanding the expression of CKX members in different plant tissues would help in understanding the specific functions of CKX members in plant tissues. In this study, the expression levels of StCKX members showed variation in different tissues, showing tissue-specific expression during the seedling stage, with certain CKX genes having higher expression levels in the leaves and roots. In contrast, others had higher expression in the stems than in other tissues. Similarly, the GmCKXs also had tissue-specific expression at the sprout stage, in which some members (such as GmCKX7, GmCKX9) were expressed in the radicle while some CKX members were expressed in the cotyledons [61]. Also, the expression of PvCKXs showed a tissue-specific feature [27], revealing that CKXs in plants might have a specific expression pattern in different tissues.
As the most sensitive growth stage of plants, the seedling stage is generally more affected by environmental changes. The expression levels of StCKX members were significantly altered during this period under stress, indicating that CKX members might be responsive to various stresses and participate in stress adaptation and resistance. Some CKX members are reportedly responsive to abiotic or biotic stresses. Specifically, the overexpression of the AtCKX3 gene in transgenic tomatoes resulted in the maintenance of a higher water state due to reduced transpiration under drought treatment and enhanced drought resistance [62]. The expression of CKX5 was significantly induced in leaves infected with Botrytis cinerea, where the overexpression of CKX5 in Arabidopsis resulted in stronger resistance to Botrytis cinerea than wild-type plants [63]. PpCKX1, expressed in vacuoles, was shown to be involved in enhancing dehydration and salt tolerance [64]. Similarly, the expression of StCKXs in this study was changed significantly under stresses (such as StCKX04, StCKX06, StCKX07, and StCKX08), which indicates that they might be responsive to stress. These results provide a theoretical basis and insights for molecular breeding in potato, with a particular focus on StCKXs.

5. Conclusions

In this study, nine StCKX members were identified in the reference genome of potato, and a comprehensive analysis was performed for their characterization, including their chromosomal location, evolutionary relationships, motifs, gene structure, cis-acting elements, collinearity, and expression patterns. The results showed that StCKXs were divided into six clusters, and the CKXs within each subgroup displayed similar characteristics. The StCKXs were also shown to likely be involved in processes related to plant hormones, stress-related factors, and sprouting. The StCKXs had tissue-specific expression patterns, and their expression levels changed when the plants were subjected to abiotic and biotic stress conditions. These observations revealed that the activity of StCKXs, the cytokinin content, and CKX expression were correlated. Finally, this study revealed valuable insights that may help engineer CKXs in potato (Solanum tuberosum) to improve crop resistance to various environmental stress and climate change.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/horticulturae10070737/s1; Table S1: The primer of StCKX members; Table S2: The function of cis-acting elements in StCKXs.

Author Contributions

Conceptualization and methodology, W.Z.; software, S.L.; formal analysis, S.W.; investigation, F.X.; resources, Z.L. and B.J.; funding acquisition, W.Z. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by Innovation Project of Heilongjiang Academy of Agricultural Sciences, grant number CX23TS24.

Data Availability Statement

The original contributions presented in the study are included in the article/Supplementary Materials, further inquiries can be directed to the corresponding authors.

Conflicts of Interest

The authors declare no conflicts of interest.

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Figure 1. The location of CKX genes along the potato genome. Only the chromosomes containing CKX genes are displayed. The chromosome length in megabases (Mb) is shown on the left-hand scale. The line colors (blue to red) represent the gene density from low to high.
Figure 1. The location of CKX genes along the potato genome. Only the chromosomes containing CKX genes are displayed. The chromosome length in megabases (Mb) is shown on the left-hand scale. The line colors (blue to red) represent the gene density from low to high.
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Figure 2. The evolutionary relationships between StCKXs. Cyan, brown, yellow, blue, purple, and pink represent clusters I–VI, respectively.
Figure 2. The evolutionary relationships between StCKXs. Cyan, brown, yellow, blue, purple, and pink represent clusters I–VI, respectively.
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Figure 3. The motifs and gene structure of StCKXs. (A) The evolutionary relationship among the nine StCKXs, with the six colors (cyan, brown, yellow, blue, purple, and pink) representing the different evolutionary clusters. (B) The motifs identified in the StCKX protein sequences were predicted by MEME. The motifs were ordered based on the calculated value. The ruler below the column represents the scale of the proteins and motifs. (C) The gene structure of the StCKXs, with the green boxes representing the CDS regions of the StCKX genes and the yellow boxes representing the UTR regions.
Figure 3. The motifs and gene structure of StCKXs. (A) The evolutionary relationship among the nine StCKXs, with the six colors (cyan, brown, yellow, blue, purple, and pink) representing the different evolutionary clusters. (B) The motifs identified in the StCKX protein sequences were predicted by MEME. The motifs were ordered based on the calculated value. The ruler below the column represents the scale of the proteins and motifs. (C) The gene structure of the StCKXs, with the green boxes representing the CDS regions of the StCKX genes and the yellow boxes representing the UTR regions.
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Figure 4. Evolutionary relationship analysis of CKXs from five plant species. The colored backgrounds represent the different evolutionary clusters. The outer ring shows the motif configurations, with the different-colored strips representing the motifs 1–10. The inner ring shows the phylogeny of CKXs members grouped in seven clusters.
Figure 4. Evolutionary relationship analysis of CKXs from five plant species. The colored backgrounds represent the different evolutionary clusters. The outer ring shows the motif configurations, with the different-colored strips representing the motifs 1–10. The inner ring shows the phylogeny of CKXs members grouped in seven clusters.
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Figure 5. The cis-acting elements in the promoter region of StCKXs. (A) Evolutionary relationships among the nine StCKXs, with the six colors representing the different StCKX clusters. (B) The cis-acting elements analysis predicted by PlantCARE, with the different shapes and colors representing the different elements.
Figure 5. The cis-acting elements in the promoter region of StCKXs. (A) Evolutionary relationships among the nine StCKXs, with the six colors representing the different StCKX clusters. (B) The cis-acting elements analysis predicted by PlantCARE, with the different shapes and colors representing the different elements.
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Figure 6. Collinearity analysis of StCKXs relative to the CKXs of other plant species. The different colored squares represent the chromosomes of different species. The red, green, and pink (AT1) chromosomes represent those of potato, soybeans, and Arabidopsis, respectively. The green lines connect the collinear pairs of CKXs with genes in soybeans, and the pink lines connect the collinear pairs with genes in Arabidopsis.
Figure 6. Collinearity analysis of StCKXs relative to the CKXs of other plant species. The different colored squares represent the chromosomes of different species. The red, green, and pink (AT1) chromosomes represent those of potato, soybeans, and Arabidopsis, respectively. The green lines connect the collinear pairs of CKXs with genes in soybeans, and the pink lines connect the collinear pairs with genes in Arabidopsis.
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Figure 7. Tissue-specific expression patterns of StCKXs at the seedling stage. The green, yellow, and brown bars represent the leaves, stems, and roots, respectively. The lowercase letters indicate significant differences (p < 0.05, Number of samples = 3).
Figure 7. Tissue-specific expression patterns of StCKXs at the seedling stage. The green, yellow, and brown bars represent the leaves, stems, and roots, respectively. The lowercase letters indicate significant differences (p < 0.05, Number of samples = 3).
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Figure 8. The tissue-specific expression patterns of StCKXs after plants were exposed to various abiotic and biotic stress conditions. The green, cyan, pink, blue, and green bars represent the control (CK), salt (S), heavy metal (H), cold (C), and Clavibacter michiganensis subsp. (Cms) treatments, respectively. The lowercase letters indicate significant differences (p < 0.05, Number of samples = 3).
Figure 8. The tissue-specific expression patterns of StCKXs after plants were exposed to various abiotic and biotic stress conditions. The green, cyan, pink, blue, and green bars represent the control (CK), salt (S), heavy metal (H), cold (C), and Clavibacter michiganensis subsp. (Cms) treatments, respectively. The lowercase letters indicate significant differences (p < 0.05, Number of samples = 3).
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Figure 9. Cytokinin oxidase activity and cytokinin content in seedlings exposed to different stress conditions. The green, cyan, pink, blue, and green bars represent the control (CK), salt (S), heavy metal (H), cold (C), and Clavibacter michiganensis subsp. (Cms) treatments, respectively. The lowercase letters indicate significant differences (p < 0.05, Number of samples = 3).
Figure 9. Cytokinin oxidase activity and cytokinin content in seedlings exposed to different stress conditions. The green, cyan, pink, blue, and green bars represent the control (CK), salt (S), heavy metal (H), cold (C), and Clavibacter michiganensis subsp. (Cms) treatments, respectively. The lowercase letters indicate significant differences (p < 0.05, Number of samples = 3).
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Table 1. Detailed characteristics of StCKX members.
Table 1. Detailed characteristics of StCKX members.
Gene NamechrLocationCDS LengthMolecular WeightTheoretical pIInstability
Index		Aliphatic
Index
StCKX01164568552-6457110638714,886.855.3336.6289.06
StCKX02411108702-1111124896335,494.135.3636.0384.72
StCKX03470221729-7022762627611,095.668.8250.160.99
StCKX04833888474-33891981162960,572.186.1434.9593.45
StCKX05833930753-33933803146455,122.16.4742.8391.01
StCKX061052575434-52575820210079,314.958.6241.491.29
StCKX071057972972-57976328118243,988.236.4233.6495.24
StCKX08123611311-3619224158159,107.356.136.0290.44
StCKX09123650404-3656647124845,612.367.1232.8697.01
Table 2. Gene ontology enrichment results for the StCKX genes. A list with the top 20 results is shown.
Table 2. Gene ontology enrichment results for the StCKX genes. A list with the top 20 results is shown.
GO_AccessionDescriptionTerm_TypeQ-Value
GO:0008762UDP-N-acetylmuramate dehydrogenase activitymolecular_function0.00
GO:0009308Amine metabolic processbiological_process0.00
GO:0009690Cytokinin metabolic processbiological_process0.00
GO:0010817Regulation of hormone levelsbiological_process0.00
GO:0016614Oxidoreductase activitymolecular_function0.00
GO:0019139Cytokinin dehydrogenase activitymolecular_function0.00
GO:0034754Cellular hormone metabolic processbiological_process0.00
GO:0042445Hormone metabolic processbiological_process0.00
GO:0048037Cofactor bindingmolecular_function0.01
GO:0050660Flavin adenine dinucleotide bindingmolecular_function0.01
GO:0050662Coenzyme bindingmolecular_function0.01
GO:0065008Regulation of biological qualitybiological_process0.01
GO:0055114Oxidation-reduction processbiological_process0.02
GO:1901564Organonitrogen compound metabolic processbiological_process0.02
GO:0016491Oxidoreductase activitymolecular_function0.02
GO:0065007Biological regulationbiological_process0.03
GO:0043168Anion bindingmolecular_function0.04
GO:0000166Nucleotide bindingmolecular_function0.04
GO:1901265Nucleoside phosphate bindingmolecular_function0.04
GO:0036094Small molecule bindingmolecular_function0.04
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Zhang, W.; Liu, S.; Wang, S.; Xu, F.; Liu, Z.; Jia, B. Cytokinin Oxidase (CKX) Family Members in Potato (Solanum tuberosum): Genome-Wide Identification and Expression Patterns at Seedling Stage under Stress. Horticulturae 2024, 10, 737. https://doi.org/10.3390/horticulturae10070737

AMA Style

Zhang W, Liu S, Wang S, Xu F, Liu Z, Jia B. Cytokinin Oxidase (CKX) Family Members in Potato (Solanum tuberosum): Genome-Wide Identification and Expression Patterns at Seedling Stage under Stress. Horticulturae. 2024; 10(7):737. https://doi.org/10.3390/horticulturae10070737

Chicago/Turabian Style

Zhang, Wei, Shangwu Liu, Shaopeng Wang, Feifei Xu, Zhenyu Liu, and Bei Jia. 2024. "Cytokinin Oxidase (CKX) Family Members in Potato (Solanum tuberosum): Genome-Wide Identification and Expression Patterns at Seedling Stage under Stress" Horticulturae 10, no. 7: 737. https://doi.org/10.3390/horticulturae10070737

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