Research article
Genome-wide characterization of two-component system (TCS) genes in melon (Cucumis melo L.)

https://doi.org/10.1016/j.plaphy.2020.03.017Get rights and content

Highlights

  • 51 TCS genes were analyzed in melon genome, including conserved motifs, domains and amino acid sequence similarity levels.

  • Segmental duplication events might be the main contributors to the expansion of melon TCS genes.

  • Numerous stress- and hormone-responsive cis-elements were detected in the putative promoter regions of melon TCS genes.

  • Melon type-A and type-B RR genes were separated based on their phylogenic relationship, gene structure and expression profile.

Abstract

To better understand cytokinin signaling in melon (Cucumis melo L.), one of the most important fruit crops in the Cucurbitaceae family, we identified and characterized melon two-component system (TCS) genes in this study. The results showed that there were 51 genes encoding putative TCS proteins in melon, and these TCS genes were classified into 3 subgroups, with 17 HK(L)s (histidine kinase/histidine-kinase like; 9 HKs and 8 HKLs), 9 HPs (histidine phosphotransfer proteins; 6 authentic and 3 pseudo), and 25 RRs (response regulators; 8 Type-A, 11 Type-B and 6 pseudo). The identity values of these cytokinin signaling proteins were revealed by analyzing their conserved motifs, domains and amino acid sequences. By analyzing TCS genes in different plant species, we found that melon HK(L)s, HPs and RRs had closer phylogenetic relationships with cucumber genes than with the genes of other plants, and the expansion of melon cytokinin signaling genes might be attributed to segmental duplication events. Analysis of the putative promoter regions (2-kb upstream regions of the start codon) revealed the enrichment of stress- and hormone-response cis-elements. The involvement of these putative TCS genes in melon cytokinin signaling was further supported by qRT-PCR data.

Introduction

Cytokinin is a key plant hormone and has been reported to be involved in almost all aspects of plant growth and development, such as seed germination, root and leaf development, flowering, and plant senescence, as well as responses to environmental stimuli (Hwang and Sheen, 2001; Werner and Schmülling, 2009; Fusconi, 2014; Han et al., 2014; Edwards et al., 2018; Kieber and Schaller, 2018). Cytokinin signaling is transduced by a two-component system (TCS) in plants. This system is first reported in bacteria and usually composed of two signaling elements, a histidine kinase (HK) gene family and a response regulator (RR) gene family (Mizuno, 1997; Stock et al., 2000). HK proteins can be phosphorylated and successively transfer the phosphoryl group to a conserved Asp residue in the receiver (Rec) domain of an RR. The phosphorylated RR protein works as a direct or indirect modulator of the activity of downstream genes (Stock et al., 2000; Hwang et al., 2012). Plant TCSs are more complex than most of bacteria TCSs, and a histidine phosphotransfer (HP) gene family has been identified to be responsible for an additional phosphorylation step (Thomason and Kay, 2000; Urao et al., 2000). HP protein functions as a bridge in the transfer of the phosphoryl group between the HK and the RR (Urao et al., 2000; Hwang et al., 2002; Schaller et al., 2008).

TCS signaling has been widely reported in plant kingdoms, such as the model plants Arabidopsis thaliana and rice (Oryza sativa), as well as in some important horticultural crops (Hwang et al., 2002; Pareek et al., 2006; Schaller et al., 2008; Ishida et al., 2009, 2010; Mochida et al., 2010; Chu et al., 2011; Liu et al., 2014; He et al., 2016a, 2016b). The fact that the TCS gene number varies greatly in different plant species indicates that high specialization of TCS members might have occurred over the course of evolution (Liu et al., 2018). Thus, systematic bioinformatics analysis of TCS members should be conducted for more plants, which could benefit our understanding of the evolution and functions of TCS signaling in plants (Koretke et al., 2000; Wuichet et al., 2010; Capra and Laub, 2012).

Melon (Cucumis melo L.) is an important member of the Cucurbitaceae family and has attracted extensive attention from horticultural scientists due to its specific biological properties (Garcia-Mas et al., 2012; Chang et al., 2017). Previous investigations of TCS genes have been performed in cucumber and watermelon, another two members in the Cucurbitaceae family, and a close association has been demonstrated between these TCS genes and some important agricultural traits (He et al., 2016a, 2016b); however, characterization of TCS genes in melon has been largely limited due to the lack of genome data. Recently, the melon genome sequencing project was completed, and its genomic data were released, providing an opportunity to perform genome-wide analysis of TCS genes in melon (Garcia-Mas et al., 2012; Ruggieri et al., 2018).

In our previous study, 44 putative TCS members were identified in melon by analyzing genome data deposited in the Cucurbit Genomics Database (http://cucurbitgenomics.org/organism/3) (Liu et al., 2018). Here, we update the information on the identification and classification of TCS genes by analyzing the improved melon genome data that were deposited in NCBI (https://www.ncbi.nlm.nih.gov/). Based on this updated TCS information, we further analyzed the characteristics of these putative melon TCS genes systematically and explored their responses to exogenous application of cytokinins by quantitative real-time PCR (qRT-PCR). Our comprehensive analysis of melon TCS genes may benefit the functional study of TCS signaling in the future.

Section snippets

Plant materials and hormone treatment

Melon (Cucumis melo L.cv. ‘Yangjiaomi’) plants were grown in the greenhouse of Shandong Agricultural University, Tai'an, Shandong Province, P.R. China from March to July 2017, and regular plant management was performed over the cultivation course. Melon seedlings at 3 weeks after germination were used to examine the expression responses of selected TCS genes to exogenous application of trans-zeatin (tZ). The treatment was performed by spraying 100 μM tZ on the second fully expanded leaves

Identification of TCS proteins in melon

Because of the publication of improved melon genome data (Ruggieri et al., 2018), we first identified putative TCS genes in melon using the same protocol described in our previous study (Liu et al., 2018). In total, 51 putative TCS members were observed in melon; notably, 7 more TCSs were identified in this study than in the previously published study (Liu et al., 2018). These TCS genes were named based on the method used in Chinese cabbage (Liu et al., 2014), cucumber and watermelon (He et

Discussion

Cucurbitaceae crops (melon, cucumber and watermelon) and Physcomitrella patens contain fewer TCS members than other plants (Huang et al., 2009; Ishida et al., 2010; He et al., 2016a; Liu et al., 2018), possibly due to their small genome size and relatively few duplication events during their evolution. It is worth mentioning that no type-C RR was found in melon (Fig. 1C, Table 1). The type-C RR is regarded as the oldest RR and might be the possible evolutionary origin of type-A RRs via

Contribution

L.W. and Z.R. conceived and designed the experiments. Q.S. provided plant material. P.L., S.W., X.W., C.C., Q.L., and C.W. performed the experiments. L.W. and P.L. analyzed the data. L.W., X.Y and R.Z. wrote the manuscript.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

This work was supported by Key R & D plan in Shandong (2018GNC110014), Prospect of Shandong Seed Project, China (2016LZGC033), ‘Taishan Scholar’ Foundation of the People's Government of Shandong Province (ts20130932), China Postdoctoral Science Foundation (2017M612741), National Natural Science Foundation of China (31501781) and Natural Science Foundation of Guangdong Province (2018A030310446).

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    These authors contribute equally to this work.

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