Lifang Hu

Genome-wide identification and phylogenetic analysis of the ERF gene family in cucumbers

Members of the ERF transcription-factor family participate in a number of biological processes, viz., responses to hormones, adaptation to biotic and abiotic stress, metabolism regulation, beneficial symbiotic interactions, cell differentiation and developmental processes. So far, no tissue-expression profile of any cucumber ERF protein has been reported in detail. Recent completion of the cucumber full-genome sequence has come to facilitate, not only genome-wide analysis of ERF family members in cucumbers themselves, but also a comparative analysis with those in Arabidopsis and rice. In this study, 103 hypothetical ERF family genes in the cucumber genome were identified, phylogenetic analysis indicating their classification into 10 groups, designated I to X. Motif analysis further indicated that most of the conserved motifs outside the AP2/ERF domain, are selectively distributed among the specific clades in the phylogenetic tree. From chromosomal localization and genome distribution analysis, it appears that tandem-duplication may have contributed to CsERF gene expansion. Intron/exon structure analysis indicated that a few CsERFs still conserved the former intron-position patterns existent in the common ancestor of monocots and eudicots. Expression analysis revealed the widespread distribution of the cucumber ERF gene family within plant tissues, thereby implying the probability of their performing various roles therein. Furthermore, members of some groups presented mutually similar expression patterns that might be related to their phylogenetic groups.

Genome-Wide Identification and Transcriptional Expression Analysis of Cucumber Superoxide Dismutase (SOD) Family in Response to Various Abiotic Stresses

Superoxide dismutase (SOD) proteins are widely present in the plant kingdom and play important roles in different biological processes. However, little is known about the SOD genes in cucumber. In this study, night SOD genes were identified from cucumber (Cucumis sativus) using bioinformatics-based methods, including 5 Cu/ZnSODs, 3 FeSODs, and 1 MnSOD. Gene structure and motif analysis indicated that most of the SOD genes have relatively conserved exon/intron arrangement and motif composition. Phylogenetic analyses with SODs from cucumber and several other species revealed that these SOD proteins can be traced back to two ancestral SODs before the divergence of monocot and dicot plants. Many cis-elements related to stress responses and plant hormones were found in the promoter sequence of each CsSOD gene. Gene expression analysis revealed that most of the CsSOD genes are expressed in almost all the tested tissues. qRT-PCR analysis of 8 selected CsSOD genes showed that these genes could respond to heat, cold, osmotic, and salt stresses. Our results provide a basis for further functional research on SOD gene family in cucumber and facilitate their potential applications in the genetic improvement of cucumber.

Molecular cloning and characterization of an ASR gene from Cucumis sativus

Abscisic acid-, stress- and ripening-induced (ASR) proteins are widely present in the plant kingdom and play important roles in different biological processes. However, no reports of ASR proteins are available in cucumber. In this study, an ASR gene (CsASR1) was identified and characterized from Cucumis sativus. CsASR1 exhibited a high content of disorder-promoting amino acids, indicating that it is an intrinsically disordered protein (IDP). CsASR1 protein was highly homologous to ASR proteins from other plant species. Expression of CsASR1 was induced by diverse abiotic stresses such as heat, PEG and NaCl, as well as by signaling molecules such as ABA and H2O2, suggesting a close relationship between CsASR1 and abiotic stress. Overexpression of CsASR1 could increase the tolerance against salinity and osmotic stress in E. coli. Transgenic Arabidopsis plants overexpressing CsASR1 exhibited higher germination rate than WT plants on MS medium containing various concentrations of NaCl. In addition, overexpression of CsASR1 in Arabidopsis resulted in significantly improved salt tolerance due to the increased activity of SOD and elevated transcripts of SOS3 and LEA4-5. Finally, CsASR1 could protect the activity of lactate dehydrogenase (LDH) from heat-induced inactivation. Taken together, our results demonstrate that CsASR1 plays an important role in abiotic stress tolerance, and it may function as an IDP to confer abiotic stress tolerance by protecting some stress-related proteins from inactivation under stress conditions.

The catalase gene family in cucumber: genome-wide identification and organization.

Abstract Catalase (CAT) is a common antioxidant enzyme in almost all living organisms. Currently, detailed reports on cucumber (Cucumis sativus L.) CAT (CsCAT) genes and tissue expression profiling are limited. In the present study, four candidate CsCAT genes were identified in cucumber. Phylogenetic analysis indicated that CsCAT1-CsCAT3 are closely related to Arabidopsis AtCAT1-AtCAT3, but no obvious counterpart was observed for CsCAT4. Intron/exon structure analysis revealed that only one of the 15 positions was completely conserved. Motif analysis showed that, unlike the CAT genes of other species, none of CsCAT genes contained all 10 motifs. Expression data showed that transcripts of all of the CsCAT genes, except CsCAT4, were detected in five tissues. Moreover, their transcription levels displayed differences under different stress treatments.

CsCAT3, a catalase gene from Cucumis sativus, confers resistance to a variety of stresses to Escherichia coli

ABSTRACT Catalase (CAT) is a key scavenging enzyme for the degradation of hydrogen peroxide (H2O2) and plays an important role in the tolerance to diverse abiotic stresses in many different organisms. In this study, we characterized the function of a catalase gene (CsCAT3) previously isolated from Cucumis sativus in the defence against a variety of stresses. Protein alignment and phylogenetic analysis revealed that CsCAT3 was clustered in the dicot group and shared 41%–95% identity with other plant CATs. Expression analyses revealed that the expression of CsCAT3 was induced by diverse abiotic stresses such as heat, polyethylene glycol, cold and NaCl treatment, as well as by signalling molecules such as abscisic acid (ABA) and H2O2. An Escherichia coli heterologous expression system was constructed to characterize the function of CsCAT3 in vitro. Its overexpression in E. coli could increase the tolerance to heat, cold, salinity and osmotic conditions. These results indicate that CsCAT3 plays important roles in abiotic stress tolerance and that CsCAT3 confers tolerance in E. coli recombinants against abiotic stresses, which may be due to the increased activities of antioxidant enzymes that reduce the oxidative damage caused by stress conditions.

Genome-wide identification of glutathione peroxidase (GPX) gene family and their response to abiotic stress in cucumber

Plant glutathione peroxidases (GPXs) are non-heme thiol peroxidases that play vital roles in maintaining H2O2 homeostasis and regulating plant response to abiotic stress. Here, we performed a comparative genomic analysis of the GPX gene family in cucumber (Cucumis sativus). As a result, a total of 6 CsGPX genes were identified, which were unevenly located in four out of the seven chromosomes in cucumber genome. Based on the phylogenetic analysis, the GPX genes of cucumber, Arabidopsis and rice could be classified into five groups. Analysis of the distribution of conserved domains of GPX proteins showed that all these proteins contain three highly conserved motifs, as well as other conserved sequences and residues. Gene structure analysis revealed a conserved exon–intron organization pattern of these genes. Through analyzing the promoter regions of CsGPX genes, many hormone-, stress-, and development-responsive cis-elements were identified. Moreover, we also investigated their expression patterns in different tissues and developmental stages as well as in response to abiotic stress and x acid (ABA) treatments. The qRT-PCR results showed that the transcripts of CsGPX genes varied largely under abiotic stress and ABA treatments at different time points. These results demonstrate that cucumber GPX gene family may function in tissue development and plant stress responses.

Molecular cloning and functional characterization of a Cu/Zn superoxide dismutase gene (CsCSD1) from Cucumis sativus

Superoxide dismutase (SOD) proteins, which are widely present in the plant kingdom, play vital roles in response to abiotic stress. However, the functions of cucumber SOD genes in response to environmental stresses remain poorly understood. In this study, a SOD gene CsCSD1 was identified and functionally characterized from cucumber (Cucumis sativus). The CsCSD1 protein was successfully expressed in E. coli, and its overexpression significantly improved the tolerance of host E. coli cells to salinity stress. Besides, overexpression of CsCSD1 enhanced salinity tolerance during germination and seedling development in transgenic Arabidopsis plants. Further analyses showed that the SOD and CAT (catalase) activities of transgenic plants were significantly higher than those of wild-type (WT) plants under normal growth conditions as well as under NaCl treatment. In addition, the expression of stress-response genes RD22, RD29B and LEA4-5 was significantly elevated in transgenic plants. Our results demonstrate that the CsCSD1 gene functions in defense against salinity stress and may be important for molecular breeding of salt-tolerant plants.

Genome-wide identification, characterization, and transcriptional analysis of the metacaspase gene family in cucumber (Cucumis sativus)

Metacaspase (MC), a family of caspase-like proteins, plays vital roles in regulating programmed cell death (PCD) during development and in response to stresses in plants. In this study, five MC genes (designated as CsMC1 to CsMC5) were identified in the cucumber (Cucumis sativus) genome. Sequence analysis revealed that CsMC1-CsMC3 belong to type I MC proteins, while CsMC4 and CsMC5 are type II MC proteins. Phylogenetic tree and conserved motif analysis of MC proteins indicated that these proteins can be classified into two groups, which are correlated with the types of these MC proteins. Gene structure analysis demonstrated that type I CsMC genes contain 4-7 introns, while all type II CsMC genes harbor one intron. In addition, many hormone-, stress-, and development-related cis-elements were identified in the promoter regions of CsMC genes. Expression analysis using RNA-seq data revealed that CsMC genes have distinct expression patterns in various tissues and developmental stages. qRT-PCR results showed that the transcript levels of CsMC genes could be regulated by various abiotic stresses such as NaCl, PEG, and cold. These results demonstrate that the cucumber MC gene family may function in tissue development and plant stress responses.

Genome-wide identification and expression analysis of YTH domain-containing RNA-binding protein family in cucumber ( Cucumis sativus )

YTH domain-containing RNA-binding proteins are involved in post-transcriptional regulation and play important roles in the growth and development as well as abiotic stress responses of plants. However, YTH genes have not been previously studied in cucumber (Cucumis sativus). In this study, a total of five YTH genes (CsYTH1–CsYTH5) were identified in cucumber, which could be mapped on three out of the seven cucumber chromosomes. All CsYTH proteins had highly conserved C-terminal YTH domains, and two of them (CsYTH1 and CsYTH4) harbored extra CCCH and P/Q/N-rich domains. The phylogenesis, conserved motifs and exon–intron structure of YTH genes from cucumber, Arabidopsis and rice were also analyzed. The phylogenetically closely clustered YTHs shared similar gene structures and conserved motifs. An analysis of the cis-acting regulatory elements in the upstream region of these genes resulted in the identification of many cis-elements related to stress, hormone and development. Expression analysis based on the transcriptome data showed that some CsYTHs had development- or tissue-specific expression. In addition, their expression levels were altered under various stresses such as salt, drought, cold, and abscisic acid (ABA) treatments. These findings lay the foundation for the functional analysis of CsYTHs in the future.

Genome-wide identification and characterization of cysteine-rich polycomb-like protein (CPP) family genes in cucumber (Cucumis sativus) and their roles in stress responses

Cysteine-rich polycomb-like protein (CPP) transcription factors are a small gene family involved in the regulation of plant growth, development and stress response. Genome-wide identification and characterization of CPP gene family have been carried out in several plant species. However, no comprehensive phylogenetic analysis or expression profiling of CPP genes in Cucumis sativus has been reported. In this study, we characterized the CPP family genes in C. sativus, and 5 CPP candidate genes were found in cucumber genome. CPP proteins from cucumber and other plant species were classified into two groups, which were further divided into five subgroups based on the phylogenetic distribution. Most CPP genes in the same subgroup shared similar gene structures and conserved motifs. Transcriptome data revealed that CsCPP genes were expressed in leaves, ovaries, flowers, stems, roots, and tendril tissues. qRT-PCR expression analysis showed that many CsCPP genes exhibited different expression patterns in cucumber leaves under abiotic stresses including salt, cold, drought, and ABA. These results demonstrate that cucumber CPP gene family may play critical roles in plant development as well as in the responses to environmental stresses.