Genome-Wide Identification and Characterization of Tea SGR Family Members Reveal Their Potential Roles in Chlorophyll Degradation and Stress Tolerance

Abstract

Photosynthesis plays vital roles in plant growth and development. Stay-green (SGR) proteins are responsible for chlorophyll degradation and photosynthetic metabolism. To identify SGR family members and determine their potential functions in tea plants, we identified and cloned three SGR genes. Phylogenetic analysis revealed that the tea SGR homologs were classified into the SGR subfamily (named CsSGR1 and CsSGR2) and the SGRL subfamily (named CsSGRL). Cis-element analysis indicated that the promoters of CsSGR1, CsSGR2 and CsSGRL contained light-, phytohormone- and stress-related elements. Subcellular localization confirmed that CsSGR1 was localized in the chloroplast, while CsSGR2 and CsSGRL were localized in the chloroplast, membrane and nucleus. The RT-qPCR results showed that the three genes in the matures of albino tea cultivars were expressed higher than in the green tea cultivar. However, only CsSGR2-overexpressing tobacco leaves exhibited a yellowish phenotype and significantly lower Fv/Fm values. CsSGR1 and CsSGR2 exhibited similar expression patterns in different tissues after infection with the pathogen Colletotrichum camelliae, which was opposite to the pattern observed for CsSGRL. In addition, CsSGR1 was significantly induced in response to cold stress, SA, JA and ABA in C. camelliae. These findings identified valuable candidate genes for elucidating the mechanism of leaf albinism, stress response and phytohormone signaling in tea plants.

1. Introduction

Chlorophylls (Chls), which mainly include Chl a and Chl b, are a class of tetrapyrrole molecules and indispensable for light energy absorption and electron transfer during photosynthesis [1]. Changes in leaf or fruit color, shifting from green to yellow or red, often coincide with the depletion of Chl [2,3]. Notably, excess Chl and its derivatives are potential cellular phytotoxins and must be rapidly degraded [4,5]. The STAY-GREEN (SGR) gene, crucial for Mg-dechelatase, plays a pivotal role in extracting central Mg from Chl a [6]. SGR1 and SGRL can interact with light-harvesting complex II (LHCII) and chlorophyll catabolic enzymes (CCEs) to form the SGR1/SGRL-CCE-LHCII complex, which accelerates Chl degradation and mitigates the damage caused by catabolites to cell membranes [2,5,7].

Armstead et al. were the first to identify the gene governing Mendel’s green cotyledon trait in Arabidopsis thaliana, Pisum sativum and Festuca pratensis [8], which is now called STAY-GREEN 1 (SGR1) or NONYELLOEING1 (NYE1). Subsequently, SGR homologs were widely identified in other plants, such as Oryza sativa [9], Solanum lycopersicum [10], Capsicum annuum [10], Medicago truncatula [11], Glycine max [12], Cucumis melo L. [13] and Citrus sinensis [14]. The SGR family is categorized into two subfamilies: the SGR and SGR-like (SGRL) subfamilies. All SGR homologs are localized in chloroplasts and contain chloroplast transit peptides, conserved SGR domains and variable C-terminal regions [2,15]. The distinguishing feature between SGR and SGRL lies in the presence of a cysteine-rich motif (CRM, C-X3C-X-C2-F-P-X5-P), essential for Mg dechelatase activity and significant in Chl degradation, which is found in the C-terminus of most SGR proteins but absent in SGRL proteins [13,15].

SGR homologs primarily participate in regulating Chl degradation, carotenoid metabolism, fruit ripening and leaf senescence [2,14,16]. Within the Arabidopsis genome, three SGR members exist as the following: SGR1/NYE1, SGR2 and SGRL. SGR1/NYE1 modulates pheophorbide a oxygenase activity to facilitate Chl degradation during plant senescence [17]. This process is regulated by the NAC016 transcription factor, which can bind to the promoter of SGR1 [7]. In contrast, the SGR2-overexpressing line remains green, and the sgr2-1 mutant displays early leaf yellowing [5]. However, Wu et al. [18] reported that SGR2 promotes Chl degradation. SGRL contributes to Chl breakdown or turnover in pre-senescent leaves [5]. In the melon, the CmSGR1- and CmSGR2-overexpressing lines exhibited fruit ripening and leaf yellowing, while the suppression of CmSGR1 and CmSGR2 delayed the degradation of Chl [13]. In the tomato, the inhibition of SlSGR1 in transgenic fruits elevated the accumulation of lycopene and β-carotene [19]. In citrus, SGRa plays a positive role in Chl degradation, and SGRa and SGRb play negative roles in carotenoid biosynthesis [14]. In addition, studies also showed that SGR genes play roles in plant resistance. A candidate SGR gene for anthracnose was identified in a Gy14 cucumber [20]. Amino acid substitution at position 108 of the SGR protein is accountable for disease resistance in the Gy14 cucumber [21]. In Medicago truncatula, the sgr mutant showed enhanced defense responses to Asian soybean rust [22].

The tea plant (Camellia sinensis (L). O. Kuntze) stands as an economically vital perennial woody species, revered for its leaves being processed into tea for drinking. Albino leaf occurrences in tea plants often stem from Chl deficiency [23]. Typically, plant leaf albinism is viewed as an aberrant phenotype leading to yield reduction and heightened vulnerability to pathogens and pests [23]. However, albino leaves often confer tea with a relatively high amino acid content and good flavor [24]. To date, several studies have revealed that SGR genes play important roles in tea leaf albinism. Transcriptional profiling revealed that the expression of the SGR gene was greater in the chlorina shoots than in green shoots of tea plants [23,25,26]. The CsSGR-CssHSP-CsLHCII complex potentially regulates the albino phenotype of tea leaves [27]. Nevertheless, the comprehensive genomic identification of SGR genes and their prospective functionalities await further elucidation.

Therefore, to identify SGR gene members and investigate their potential roles in tea plants, the genome-wide identification and cloning of CsSGRs was conducted in this study. The evolutionary relationships, protein characteristics, subcellular localizations, expression patterns and functions of CsSGRs in Chl degradation were analyzed. Our study identified CsSGR genes in tea plants and provides valuable insights into their potential roles. The results will lay the foundation for further functional elucidation of CsSGR members, which will be helpful for breeding elite tea cultivars with albino or other desirable phenotypes.

2. Materials and Methods

2.1. Plant Materials and Treatments

Tissues from various parts of tea plants, including the buds, leaves from different positions (1st, 2nd, 3rd and 4th leaves) and stems, were gathered for RT-qPCR analysis from 5-year-old ‘Longjing 43’ (LJ43) tea plants cultivated at the Tea Research Institute of the Chinese Academy of Agricultural Science in Hangzhou, China. Given the association of SGR genes with Chl degradation during leaf senescence, mature leaves from a green tea cultivar (‘Longjing43’), light-sensitive albino tea cultivars (‘Zhonghuang 1’ and ‘Huangjinya’) and a temperature-sensitive albino tea cultivar (‘Baiye 1’) were collected for RT-qPCR analysis.

For biotic stress induction, 5-year-old ‘LJ43’ tea plants, grown in a greenhouse for a month, were subjected to acupuncture and inoculated with a conidial suspension of C. camelliae, as previously described [28]. Samples were gathered at 0, 6, 12, 24, 48 and 72 h post-treatment, while a mock group was inoculated with sterile water. For cold treatment, the ‘LJ43’ tea plants were exposed to either 4 °C or 22 °C. Young leaves were collected at 6, 12, 24 and 48 h post-treatment. Regarding jasmonic acid (JA), salicylic acid (SA) and abascisic acid (ABA) treatments, detached branches of ‘LJ43’ were sprayed with 150 μg mL⁻¹ JA, 1 mM SA and 100 μM ABA [29]. One bud and one leaf were collected at 1, 3, 6, 12 and 24 h post-treatment, with samples sprayed with sterile water serving as controls. Each experiment utilized three independent biological replicates.

2.2. Identification and Bioinformatics Analysis of SGR Genes in Tea Plants

All sequences pertaining to tea plants were sourced from the Tea Plant Information Archive (TPIA, http://tpia.teaplants.cn/index.html, accessed on 4 November 2023). SGR protein sequences from Arabidopsis thaliana and Solanum lycopersicum [2] were queried via the TPIA online tool with BLASTp (e-value ≤ 1 × 10⁻¹⁰) against the ‘Longjing 43’, ‘Shuchazao’ and ‘Tieguanyin’ databases to pinpoint tea plant CsSGR genes. Furthermore, the Pfam database (http://pfam.xfam.org, accessed on 4 November 2023) and the Conserved Domain Database (CDD, https://www.ncbi.nlm.nih.gov/cdd, accessed on 4 November 2023) were utilized to verify that the identified genes harbored a stay-green domain. Additionally, the conserved motifs within CsSGR proteins were determined using the MEME database (https://meme-suite.org/meme/, accessed on 4 November 2023). A phylogenetic tree was constructed using MEGA version 11 software employing the neighbor-joining method with 1000 bootstrap trials. Multiple sequence alignment of SGR proteins was conducted using DNAMAN. Lastly, the PlantCARE database (http://bioinformatics.psb.ugent.be/webtools/plantcare/html/, accessed on 7 November 2023) was employed to analyze cis-acting regulatory elements within the promoters.

2.3. Cloning and RT-qPCR of CsSGR1, CsSGR2 and CsSGRL

All samples were promptly frozen in liquid nitrogen, followed by storage at −80 °C for subsequent analyses. Total RNA extraction from tea samples was performed using a FastPure Plant Total RNA Isolation Kit (Polysaccharides & Polyphenolics-Rich) (Nanjing Vazyme Biotech Co., Ltd., Nanjing, China) according to the manufacturer’s instructions. Subsequently, 1 μg of total RNA was utilized for first-strand cDNA synthesis by employing the HiScript II 1st Strand cDNA Synthesis Kit (with gDNA wiper) (Nanjing Vazyme Biotech Co., Ltd., Nanjing, China). The cDNA from ‘LJ43’ leaves served as a template for amplifying the CDS of CsSGR1, CsSGR2 and CsSGRL using Phanta^® Max ultra-fidelity DNA polymerase (Nanjing Vazyme Biotech Co., Ltd., Nanjing, China). RT-qPCR was performed following established protocols [30,31], with the CsPTB reference gene serving as the internal control [32]. The primer sequences utilized for gene cloning and RT-qPCR are detailed in Supplementary Table S1.

2.4. Subcellular Localization of CsSGR1, CsSGR2 and CsSGR

To elucidate the subcellular localization of CsSGR1, CsSGR2 and CsSGRL, their ORFs lacking stop codons were integrated into the pCAMBIA2300-35S::GFP vector, resulting in the 35S::CsSGR1-GFP, 35S::CsSGR2-GFP and 35S::CsSGRL-GFP vectors. These recombinant vector plasmids were subsequently introduced into the Agrobacterium strain GV3101 and transiently expressed in Nicotiana benthamiana leaves. After 48 h, fluorescence signals were assessed utilizing a laser scanning confocal microscope (LSM 980; Zeiss, Oberkochen, Germany). The primers employed for constructing recombinant vectors are detailed in Supplementary Table S1.

2.5. Measurement of Fv/Fm

The subcellular localization vectors were used for transient overexpression analysis. The entire leaves of Nicotiana benthamiana were inoculated with the pCAMBIA2300, 35S::CsSGR1-GFP, 35S::CsSGR2-GFP and 35S::CsSGRL-GFP vectors via the Agrobacterium tumefaciens-mediated method. After 48 h, the inoculated plants were subjected to a 20 min dark period, following which the Fv/Fm of the transformed leaves was measured using a FluorCam 7 (Photon Systems Instruments, Banbury, UK) [33]. The PAM mode was used for Fv/Fm analysis. For Fo measurement, the duration was set as 5.04 s. For Fm measurement, the pulse duration and intensity were set as 0.8 s and 20%, respectively. Each treatment was conducted with nine independent biological replicates.

2.6. Statistical Analysis

The data in this study are depicted as the means ± SEMs of at least three biological independent replicates. Statistical analyses were conducted using SPSS version 27 software, employing ANOVA followed by the LSD test or t-test. Statistical significance is denoted by asterisks (* p < 0.05 and ** p < 0.01). Figures were generated using GraphPad Prism 8 and Adobe Photoshop 23.0.

3. Results

3.1. Identification, Cloning and Evolutionary Analysis of CsSGRs in Tea Plants

In this study, a total of six, putative, full-length genes encoding stay-green proteins were identified via BLASTp searches of AtSGR and SlSGR protein sequences against the tea plant genome and verified via CDD and MEME analyses. Three of the six CsSGRs were classified into the SGR subfamily, and TEA010303 and GWHPASIV042313 shared 100% similarity in protein sequences. The other three CsSGRs were classified into the SGRL subfamily, and GWHTACFB016019 and GWHTASIV012516 shared 94.12% similarity in protein sequences (Supplementary Figure S1; Supplementary Table S2). According to these reference sequences, multiple specific primers were designed and RT-PCR was performed using cDNA from the ‘Longjing 43’ tea cultivar as a template to clone CsSGR genes. Finally, three CsSGR genes were obtained and their protein sequences were used to construct a phylogenetic tree to explore the evolutionary relationships of CsSGR homologs. The two CsSGR proteins that were most closely related to VvSGR1 were classified into the SGR subfamily and named CsSGR1 and CsSGR2. One protein that shared the highest homology with SlSGRL and NtSGRL was divided into the SGRL subfamily and named CsSGRL (Figure 1A).

3.2. Sequence and Conserved Element Analysis of CsSGR1, CsSGR2 and CsSGRL in Tea Plants

To explore the structural features and functional differentiation of CsSGR1, CsSGR2 and CsSGRL, we analyzed their sequences and conserved elements. CsSGR1, CsSGR2 and CsSGRL consisted of 216, 286 and 260 amino acid residues, respectively (Figure 2). All three members contained a chloroplast transmit peptide, an SGR domain and a variable C-terminal region (Figure 1B, Figure 2). Conserved motif analysis indicated that CsSGR2 possesses a cysteine-rich motif (CRM) in the C-terminus (Figure 1C–motif 9, Figure 2), which is crucial for Mg dechelatase-mediated Chl degradation [2,15], but this motif was absent in CsSGR1 or CsSGRL. These results suggest that CsSGR2 is more likely to participate in Chl degradation.

3.3. Cis Element Analysis of CsSGR Promoters

The cis-acting regulatory elements were analyzed using 2000 bp promoter sequences upstream of the coding region to explore the potential functions of CsSGR1, CsSGR2 and CsSGRL. The results showed that light- and phytohormone-responsive elements were present in CsSGR1 (light-, auxin-, salicylic acid-, MeJA- and gibberellin-responsive elements), CsSGR2 (light-, abscisic acid- and gibberellin-responsive elements) and CsSGRL (light-, MeJA- and abscisic acid-responsive elements). In addition, stress-related responsive elements such as low-temperature, defense and anaerobic elements were present in the CsSGR1 promoter, while anoxic-specific inducibility elements were present in the CsSGR2 promoter, and defense elements were present in the CsSGRL promoter (Figure 1D; Supplementary Table S3). These results suggest that in addition to being involved in photosynthetic processes, CsSGR1, CsSGR2 and CsSGRL might be involved in stress response processes and participate in phytohormone signaling pathways.

3.4. Subcellular Localization of CsSGR1, CsSGR2 and CsSGRL

To investigate the subcellular localizations and potential functions of CsSGR1, CsSGR2 and CsSGRL, their coding sequences without stop codons were inserted into the pCAMBIA2300-35S::GFP vector. The resulting recombinant plasmids, namely 35S::CsSGR1-GFP, 35S::CsSGR2-GFP and 35S::CsSGRL-GFP, were transiently expressed in H2B-RFP transgenic N. benthamiana leaves via Agrobacterium tumefaciens-mediated transformation [34]. The GFP fluorescence signal of the vector control (35S::GFP) was observed throughout the cell. CsSGR1, CsSGR2 and CsSGRL were localized in the chloroplast. In addition, CsSGR2 and CsSGRL were also detected in the membrane and nucleus (Figure 3). These results further imply the potential functions of CsSGR1, CsSGR2 and CsSGRL in Chl degradation or leaf photosynthesis.

3.5. Expression Patterns of CsSGR1, CsSGR2 and CsSGRL in Different Tea Cultivars and Tissues

The SGR genes play crucial role in Chl degradation, closely linked to the leaf albinism in tea plants. Consequently, the expression levels of CsSGR1, CsSGR2 and CsSGRL were examined in the mature leaves of the green tea cultivar (‘Longjing43’) and light-sensitive albino tea cultivars (‘Zhonghuang 1’ and ‘Huangjinya’), as well as the temperature-sensitive albino tea cultivar (‘Baiye 1’). The results showed that compared with those in ‘Longjing 43’, the expression patterns of CsSGR1, CsSGR2 and CsSGRL were similar and were greatest in the mature leaves of ‘Baiye 1’, followed by ‘Huangjinya’ and ‘Zhonghuang 1’ mature leaves (Figure 4A,B), suggesting that these genes may play important roles in the albinism of tea leaves.

Furthermore, the expression patterns of CsSGR1, CsSGR2 and CsSGRL were examined across various tissues of tea plants. The RT-qPCR results revealed that the expression patterns of CsSGR1 and CsSGR2 were similar, contrasting with that of CsSGRL. CsSGR1 and CsSGR2 were highly expressed in the stem (S), followed by the first leaf (L1), the fourth leaf (L4), the second leaf (L2) and the third leaf (L3), while those of CsSGR2 were S > L1 > L4 > L3 > L2. CsSGRL was highly expressed in the tender tissues, including the L1 and L2, followed by the L3, bud, L4 and stem (Figure 4C). These findings suggest that the three genes may fulfill distinct roles in the development of various tissues.

3.6. Expression Patterns of CsSGR1, CsSGR2 and CsSGRL under Various Stresses and Phytohormone Treatments

Stress-related and phytohormone-responsive elements were detected in the promoters of CsSGR1, CsSGR2 and CsSGRL. To determine the potential functions of the three genes, RT-qPCR analyses were performed on tea plants subjected to biotic stress (C. camelliae), abiotic stress (low temperature) and phytohormones (SA, JA and ABA) treatment (Figure 5). CsSGR1 and CsSGR2 were significantly induced by C. camelliae infection and by JA and ABA treatment. In addition, CsSGR1 was also significantly induced at 48 h under 4 °C treatment, while the expression of CsSGR2 was repressed at 6, 12 and 24 h and increased at 48 h under 4 °C treatment. The expression of the CsSGRL gene significantly increased at 6 and 12 h but decreased at 24, 48 and 72 h after treatment with C. camelliae. There were no significant changes in the expression of CsSGRL under low-temperature or phytohormone treatments. These results indicate that CsSGR1 and CsSGR2 might play roles in the response to biotic and abiotic stresses and that phytohormones may be involved in these processes.

3.7. Functions of CsSGR1, CsSGR2 and CsSGRL in Chlorophyll Degradation

To determine the functions of CsSGR1, CsSGR2 and CsSGRL in Chl degradation, transient overexpression of CsSGR1, CsSGR2 and CsSGRL was performed in tobacco leaves. Compared with those of a wild type, pCAMBIA2300, 35S::CsSGR1 and 35S::CsSGRL leaves, 35S::CsSGR2 tobacco leaves exhibited a yellowish phenotype and significantly lower Fv/Fm values (Figure 6). Notably, compared with CsSGR1 and CsSGRL, CsSGR2 plays important roles in Chl degradation.

4. Discussion

SGR homologs play important roles in Chl degradation during leaf senescence and fruit ripening. In recent years, several studies have revealed that SGR genes are involved in leaf albinism in tea plants via transcriptomic profiling. However, comprehensive studies on SGR genes in C. sinensis are lacking. In our study, we screened and cloned three SGR members in tea plants. Phylogenetic analysis indicated that these genes could be divided into the SGR and SGRL subfamilies, which were named CsSGR1, CsSGR2 and CsSGRL, respectively. All members were found to localize in chloroplasts and featured a chloroplast transit peptide, an SGR domain and a variable C-terminal region, consistent with observations in other plant species [2,15]. Notably, the cys-rich motif (CRM, C-X3-C-X-C2-F-P-X5-P), crucial for Mg-dechelatase activity, is commonly present in most SGR proteins but absent in SGRL proteins [2]. Our MEME and multiple sequence alignment results revealed that the CRM motif was present in the CsSGR2 protein but not in the CsSGR1 and CsSGRL proteins. The nonexistence of the CRM motif was also found in the CmSGR3 and CmSGR4 proteins of Cucumis melo L. [13] and the GmaSGR4 protein of Glycine max [2]. Due to the Chl degradation function of CRM, CsSGR2 may have a greater ability to mediate Chl degradation than CsSGR1 and CsSGRL. The 35S::CsSGR2 tobacco leaves exhibited a yellowish phenotype and significantly lower Fv/Fm values in our study, further suggesting that the CRM motif is important for Chl degradation. Notably, although the CRM motif is absent in SGR or SGRL proteins, these proteins may be recruited or activated by maturity, stress or phytohormones to mediate Chl degradation [35]; however, this mechanism needs to be further studied in tea plants. In addition, SGR and SGRL may function at different stages of plant development in Arabidopsis, SGR1 functions in Chl degradation during natural senescence and the expression level of SGR1 mainly increases with tissue aging, and SGRL may play a vital role in Chl turnover under pre-senescence conditions with a high expression level in green leaves [2]. The expression patterns of CsSGR and CsSGRL in tea plants are similar to those in Arabidopsis, indicating that CsSGR and CsSGRL may play different roles in Chl degradation at different stages of tea senescence.

Traditionally, SGR proteins are responsible for the extraction of central Mg from Chl a, which results in Chl degradation. CRM, which participates in inter- or intramolecular crosslinking or redox regulation, is responsible for Chl degradation [15]. However, the presence or absence of CRM results in different functions in different species. Early termination of SGR led to a lack of CRM and resulted in a stay-green trait in pak choi [36,37]. In citrus, the overexpression of truncated SGRa^stop resulted in no obvious Chl degradation [14]. Interestingly, we found that the length of CsSGR1 (216 amino acid residues) in tea plants was shorter than that of most other SGR proteins, and the CRM motif was also absent in CsSGR1. It may be that the truncated version of SGR possesses different functions compared with traditional SGR homologs. Although our study revealed significantly higher expression of CsSGR1 in ‘Huangjinya’ and ‘Baiye 1’ leaves compared to ‘Longjing 43’ leaves, the 35S::CsSGR1 tobacco leaves displayed a phenotype similar to that of the wild-type plants. These findings suggest that CsSGR1 may not have a decisive role in Chl degradation, and the absence of CRM deficiency might contribute to this observation.

In addition to functioning in promoting Chl degradation by SGR homologs, recent studies have shown that they participate in biotic and abiotic stress resistance via the Chl degradation pathway. The central Mg of Chl a is extracted by Mg-dechelatase encoded by SGR1 to form pheophytin a [6], which is subsequently catalyzed to pheophorbide a (Pheide a) and red Chl catabolite (RCC) [35]. Pheide a and RCC, which can potentially be phototoxic, have the ability to induce cell death or generate ROS (especially H₂O₂) which function as signaling molecules to mediate defense responses against pathogenic infection [21,35,38]. In addition, SGR can interact with chlorophyll’s catabolic enzymes (CCEs) and light-harvesting complex II (LHCII) to form the SGR-CCE-LHCII complex, contributing to the detoxification of phototoxic catabolites [2]. In Cucumis sativus and Medicago truncatula, the mutation or silencing of SGR is responsible for anthracnose resistance by regulating the level of ROS and the expression of defense-related genes [20,21,22]. LpSGR-mediated Chl a catabolism is involved in ryegrass heat stress tolerance [39]. In addition, phytohormones such as ABA are involved in SGR-mediated Chl breakdown and other processes [2,30]. Cis-element analysis of the promoters revealed that phytohormones and stress-related responsive elements were present in the promoters of CsSGR1, CsSGR2 and CsSGRL. Our RT-qPCR results indicated that the expression level of CsSGR1 was significantly increased in response to treatment with C. camelliae, low temperature, SA, JA and ABA. The abundance of CsSGR2 increased under treatment with C. camelliae, JA and ABA. However, the expression of CsSGRL significantly decreased in response to C. camelliae, which was opposite to what was observed for CsSGR1 and CsSGR2. These findings suggest that CsSGR genes, especially CsSGR1, may play crucial roles in stress-related processes and phytohormone signaling pathways, which needs to be further investigated. Our results also found that the CsSGR genes were expressed higher in the mature leaves of albino tea cultivars than those of the green tea cultivar. However, the resistance of the tea plant to stress is determined by many factors. Defects in chloroplast development and lower contents of secondary metabolites such as catechins and anthocyanins often present in albino tea cultivars may affect their stress tolerance [23,40]. Thus, the exact resistance function of CsSGRs in different tea cultivars remains to be clarified.

5. Conclusions

CsSGR1, CsSGR2 and CsSGRL were identified and cloned from tea plants. The promoter of the three genes contained light-, phytohormone- and stress-related elements. CsSGR1 was localized in the chloroplast, while CsSGR2 and CsSGRL were localized in the chloroplast, membrane and nucleus. RT-qPCR showed that the expression patterns of CsSGR1 and CsSGR2 were increased with tissue aging, which was opposite to CsSGRL. The expression levels of the three genes were greater in the albino tea cultivars than in the green tea cultivar. However, only CsSGR2-overexpressing tobacco leaves exhibited a yellowish phenotype and significantly lower Fv/Fm values. In addition, CsSGR1 was significantly induced in response to C. camelliae, cold stress, SA, JA and ABA treatments. This study provides candidate genes for the further functional elucidation of the mechanism of CsSGRs in the leaf albinism, stress response and phytohormone signaling of tea plants.