scholarly journals Identification and analysis of CYP450 supergene family members from the transcriptome of Aralia elata (Miq.) Seem reveal candidate genes for triterpenoid saponin biosynthesis

2020 ◽  
Author(s):  
Yao Cheng ◽  
Hanbing Liu ◽  
Xuejiao Tong ◽  
Zaimin Liu ◽  
Xin Zhang ◽  
...  
Keyword(s):  
Gene Family ◽  
Candidate Genes ◽  
Expression Patterns ◽  
Family Members ◽  
Full Length ◽  
Sequence Information ◽  
Triterpenoid Saponin ◽  
Triterpenoid Saponins ◽  
Qrt Pcr ◽  
Aralia Elata

Abstract Background: Members of the cytochrome P450 (CYP450) gene superfamily have been shown to play essential roles in regulating secondary metabolites biosynthesis. However, the systematic identification and bioinformatics analysis of CYP450s have not been reported in Aralia elata (Miq.) Seem , a highly valued medicinal plant. Results: In the present study we conducted the RNA-sequencing (RNA-seq) analysis of the leaves, stems, and roots of A. elata, yielding 66,713 total unigenes. Following the annotation and classification of these unigenes, we were able to identify two pathways and 19 putative genes associated with the synthesis of triterpenoid saponins in these plants, with qRT-PCR subsequently being used to validate these gene expression patterns. Scanning with the CYP450 model from Pfam resulted in the identification of 111 full-length and 143 partial-length CYP450s, with the full-length CYP450s being further clustered into 7 clans and 36 families. Through phylogenetic and conserved motif analyses, we were further able to group these CYP450 proteins into two primary branches: A-type (53%) and non-A type (47%). We further conducted representative protein sequence alignment for these CYP450 family members, with secondary elements being assigned in light of the recently published Arabidopsis CYP90B1 structure. Using the available sequence information, we further identified predicted substrate recognition sites (SRSs) and substrate binding sites within these putative proteins.We further assessed the expression patterns of these 111 CYP450 genes across A. elata tissues, with 12 members of this gene family being selected at random for qRT-PCR validation. From these data, we identified CYP716A295 and CYP716A296 as the candidate genes most likely to be associated with oleanolic acid synthesis, while CYP72A763 was identified as being the most likely to play a role in hederagenin biosynthesis. Finally, we assessed the subcellular localization of these CYP450 proteins within Arabidopsis protoplasts, highlighting the fact that they localize to the endoplasmic reticulum. Conclusions: This study presents a systematic analysis of the CYP450 gene family in A. elata and provided a foundation for further functional characterization of CYP450 genes.

scholarly journals Identification and analysis of CYP450 supergene family members from the transcriptome of Aralia elata (Miq.) Seem reveal candidate genes for triterpenoid saponins biosynthesis

2020 ◽  
Author(s):  
Yao Cheng ◽  
Hanbing Liu ◽  
Xuejiao Tong ◽  
Xin Zhang ◽  
Dalong Li ◽  
...  
Keyword(s):  
Gene Family ◽  
Candidate Genes ◽  
Expression Patterns ◽  
Functional Characterization ◽  
Family Members ◽  
Full Length ◽  
Sequence Information ◽  
Triterpenoid Saponins ◽  
Qrt Pcr ◽  
Aralia Elata

Abstract BackgroundMembers of the cytochrome P450 (CYP450) gene superfamily have been shown to play essential roles in regulating secondary metabolites biosynthesis. However, the systematic identification and bioinformatics analysis of CYP450s have not been reported in Aralia elata (Miq.) Seem, a highly valued medicinal plant. ResultsIn the present study we conducted the RNA-sequencing (RNA-seq) analysis of the leaves, stems, and roots of A. elata, yielding 66,713 total unigenes. Following the annotation and classification of these unigenes, we were able to identify two pathways and 19 putative genes associated with the synthesis of triterpenoid saponins in these plants, with qRT-PCR subsequently being used to validate these gene expression patterns. Scanning with the CYP450 model from Pfam resulted in the identification of 111 full-length and 143 partial-length CYP450s, with the full-length CYP450s being further clustered into 7 clans and 36 families. Through phylogenetic and conserved motif analyses, we were further able to group these CYP450 proteins into two primary branches: A-type (53%) and non-A type (47%). We further conducted representative protein sequence alignment for these CYP450 family members, with secondary elements being assigned in light of the recently published Arabidopsis CYP90B1 structure. Using the available sequence information, we further identified predicted substrate recognition sites (SRSs) and substrate binding sites within these putative proteins.We further assessed the expression patterns of these 111 CYP450 genes across A. elata tissues, with 12 members of this gene family being selected at random for qRT-PCR validation. From these data, we identified CYP716A295 and CYP716A296 as the candidate genes most likely to be associated with oleanolic acid synthesis, while CYP72A763 was identified as being the most likely to play a role in hederagenin biosynthesis. Finally, we assessed the subcellular localization of these CYP450 proteins within Arabidopsis protoplasts, highlighting the fact that they localize to the endoplasmic reticulum.ConclusionsThis study presents a systematic analysis of the CYP450 gene family in A. elata and provided a foundation for further functional characterization of CYP450 genes.

scholarly journals Identification and analysis of CYP450 and UGT supergene family members from the transcriptome of Aralia elata (Miq.) Seem reveal candidate genes for triterpenoid saponin biosynthesis

2020 ◽  
Author(s):  
Yao Cheng ◽  
Hanbing Liu ◽  
Xuejiao Tong ◽  
Zaimin Liu ◽  
Xin Zhang ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Oleanolic Acid ◽  
Expression Patterns ◽  
Functional Characterization ◽  
Acid Synthesis ◽  
Triterpenoid Saponin ◽  
Specific Expression ◽  
Systematic Analysis ◽  
Saponin Biosynthesis ◽  
Aralia Elata

Abstract Background: Members of the cytochrome P450 (CYP450) and UDP-glycosyltransferases (UGT) gene superfamily have been shown to play essential roles in regulating secondary metabolites biosynthesis. However, the systematic identification of CYP450s and UGTs have not been reported in Aralia elata (Miq.) Seem , a highly valued medicinal plant. Results: In the present study we conducted the RNA-sequencing (RNA-seq) analysis of the leaves, stems, and roots of A. elata, yielding 66,713 total unigenes. Following the annotation and KEGG pathway analysis, we were able to identify 64 unigenes related to triterpenoid skeleton biosynthesis, 254 CYP450s and 122 UGTs, respectively. 150 CYP450s and 92 UGTs encoding >300 amino acid proteins were utilized for phylogenetic and tissue-specific expression analyses. This allowed us to cluster 150 CYP450s into 9 clans and 40 families, and then these CYP450 proteins were further grouped into two primary branches: A-type (53%) and non-A type (47%). A phylogenetic analysis of 92 UGTs and other plant UGTs led to clustering into 16 groups (A-P). We further assessed the expression patterns of these CYP450 and UGT genes across A. elata tissues, with 23 CYP450 and 16 UGT members being selected for qRT-PCR validation, respectively. From these data, we identified CYP716A295 and CYP716A296 as the candidate genes most likely to be associated with oleanolic acid synthesis, while CYP72A763 and CYP72A776 was identified as being the most likely to play a role in hederagenin biosynthesis. We also selected five unigenes as the best candidates for oleanolic acid 3-O-glucosyltransferase. Finally, we assessed the subcellular localization of three CYP450 proteins within Arabidopsis protoplasts, highlighting the fact that they localize to the endoplasmic reticulum. Conclusions: This study presents a systematic analysis of the CYP450 and UGT gene family in A. elata and provided a foundation for further functional characterization of these two multigene family.

scholarly journals Identification and analysis of CYP450 and UGT supergene family members from the transcriptome of Aralia elata (Miq.) Seem reveal candidate genes for triterpenoid saponin biosynthesis

2020 ◽  
Author(s):  
Yao Cheng ◽  
Hanbing Liu ◽  
Xuejiao Tong ◽  
Zaimin Liu ◽  
Xin Zhang ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Oleanolic Acid ◽  
Expression Patterns ◽  
Functional Characterization ◽  
Acid Synthesis ◽  
Triterpenoid Saponin ◽  
Specific Expression ◽  
Systematic Analysis ◽  
Saponin Biosynthesis ◽  
Aralia Elata

Abstract Background: Members of the cytochrome P450 (CYP450) and UDP-glycosyltransferase (UGT) gene superfamily have been shown to play essential roles in regulating secondary metabolite biosynthesis. However, the systematic identification of CYP450s and UGTs has not been reported in Aralia elata (Miq.) Seem, a highly valued medicinal plant. Results: In the present study, we conducted the RNA-sequencing (RNA-seq) analysis of the leaves, stems, and roots of A. elata, yielding 66,713 total unigenes. Following annotation and KEGG pathway analysis, we were able to identify 64 unigenes related to triterpenoid skeleton biosynthesis, 254 CYP450s and 122 UGTs, respectively. A total of 150 CYP450s and 92 UGTs encoding >300 amino acid proteins were utilized for phylogenetic and tissue-specific expression analyses. This allowed us to cluster 150 CYP450s into 9 clans and 40 families, and then these CYP450 proteins were further grouped into two primary branches: A-type (53%) and non-A-type (47%). A phylogenetic analysis of 92 UGTs and other plant UGTs led to clustering into 16 groups (A-P). We further assessed the expression patterns of these CYP450 and UGT genes across A. elata tissues, with 23 CYP450 and 16 UGT members being selected for qRT-PCR validation, respectively. From these data, we identified CYP716A295 and CYP716A296 as the candidate genes most likely to be associated with oleanolic acid synthesis, while CYP72A763 and CYP72A776 were identified as being the most likely to play roles in hederagenin biosynthesis. We also selected five unigenes as the best candidates for oleanolic acid 3-O-glucosyltransferase. Finally, we assessed the subcellular localization of three CYP450 proteins within Arabidopsis protoplasts, highlighting the fact that they localize to the endoplasmic reticulum.Conclusions: This study presents a systematic analysis of the CYP450 and UGT gene family in A. elata and provides a foundation for further functional characterization of these two multigene families.

scholarly journals Identification and analysis of CYP450 and UGT supergene family members from the transcriptome of Aralia elata (Miq.) Seem reveal candidate genes for triterpenoid saponin biosynthesis

2020 ◽  
Author(s):  
Yao Cheng ◽  
Hanbing Liu ◽  
Xuejiao Tong ◽  
Zaimin Liu ◽  
Xin Zhang ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Oleanolic Acid ◽  
Expression Patterns ◽  
Functional Characterization ◽  
Acid Synthesis ◽  
Triterpenoid Saponin ◽  
Specific Expression ◽  
Systematic Analysis ◽  
Saponin Biosynthesis ◽  
Aralia Elata

Abstract Background: Members of the cytochrome P450 (CYP450) and UDP-glycosyltransferase (UGT) gene superfamily have been shown to play essential roles in regulating secondary metabolite biosynthesis. However, the systematic identification of CYP450s and UGTs has not been reported in Aralia elata (Miq.) Seem, a highly valued medicinal plant. Results: In the present study, we conducted the RNA-sequencing (RNA-seq) analysis of the leaves, stems, and roots of A. elata, yielding 66,713 total unigenes. Following annotation and KEGG pathway analysis, we were able to identify 64 unigenes related to triterpenoid skeleton biosynthesis, 254 CYP450s and 122 UGTs, respectively. A total of 150 CYP450s and 92 UGTs encoding >300 amino acid proteins were utilized for phylogenetic and tissue-specific expression analyses. This allowed us to cluster 150 CYP450s into 9 clans and 40 families, and then these CYP450 proteins were further grouped into two primary branches: A-type (53%) and non-A-type (47%). A phylogenetic analysis of 92 UGTs and other plant UGTs led to clustering into 16 groups (A-P). We further assessed the expression patterns of these CYP450 and UGT genes across A. elata tissues, with 23 CYP450 and 16 UGT members being selected for qRT-PCR validation, respectively. From these data, we identified CYP716A295 and CYP716A296 as the candidate genes most likely to be associated with oleanolic acid synthesis, while CYP72A763 and CYP72A776 were identified as being the most likely to play roles in hederagenin biosynthesis. We also selected five unigenes as the best candidates for oleanolic acid 3-O-glucosyltransferase. Finally, we assessed the subcellular localization of three CYP450 proteins within Arabidopsis protoplasts, highlighting the fact that they localize to the endoplasmic reticulum.Conclusions: This study presents a systematic analysis of the CYP450 and UGT gene family in A. elata and provides a foundation for further functional characterization of these two multigene families.

scholarly journals Identification, Evolutionary and Expression Analysis of PYL-PP2C-SnRK2s Gene Families in Soybean

Plants ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 1356
Author(s):  
Zhaohan Zhang ◽  
Shahid Ali ◽  
Tianxu Zhang ◽  
Wanpeng Wang ◽  
Linan Xie
Keyword(s):  
Gene Family ◽  
Higher Plants ◽  
Expression Patterns ◽  
Family Members ◽  
Evolutionary Relationship ◽  
Gene Families ◽  
Class Iii ◽  
Sequencing Data ◽  
Entire Genome ◽  
Core Components

Abscisic acid (ABA) plays a crucial role in various aspects of plant growth and development, including fruit development and ripening, seed dormancy, and involvement in response to various environmental stresses. In almost all higher plants, ABA signal transduction requires three core components; namely, PYR/PYL/RCAR ABA receptors (PYLs), type 2C protein phosphatases (PP2Cs), and class III SNF-1-related protein kinase 2 (SnRK2s). The exploration of these three core components is not comprehensive in soybean. This study identified the GmPYL-PP2C-SnRK2s gene family members by using the JGI Phytozome and NCBI database. The gene family composition, conservation, gene structure, evolutionary relationship, cis-acting elements of promoter regions, and its coding protein domains were analyzed. In the entire genome of the soybean, there are 21 PYLs, 36 PP2Cs, and 21 SnRK2s genes; further, by phylogenetic and conservation analysis, 21 PYLs genes are classified into 3 groups, 36 PP2Cs genes are classified into seven groups, and 21 SnRK2s genes are classified into 3 groups. The conserved motifs and domain analysis showed that all the GmPYLs gene family members contain START-like domains, the GmPP2Cs gene family contains PP2Cc domains, and the GmSnRK2s gene family contains S_TK domains, respectively. Furthermore, based on the high-throughput transcriptome sequencing data, the results showed differences in the expression patterns of GmPYL-PP2C-SnRK2s gene families in different tissue parts of the same variety, and the same tissue part of different varieties. Our study provides a basis for further elucidation of the identification of GmPYL-PP2C-SnRK2s gene family members and analysis of their evolution and expression patterns, which helps to understand the molecular mechanism of soybean response to abiotic stress. In addition, this provides a conceptual basis for future studies of the soybean ABA core signal pathway.

Identification and genes expression analysis of ATP-dependent phosphofructokinase family members among three Saccharum species

2013 ◽  
Vol 40 (4) ◽  
pp. 369 ◽  
Author(s):  
Lin Zhu ◽  
Jisen Zhang ◽  
Youqiang Chen ◽  
Hongyu Pan ◽  
Ray Ming
Keyword(s):  
Gene Family ◽  
Expression Patterns ◽  
Family Members ◽  
Sucrose Metabolism ◽  
Expression Level ◽  
Group Iii ◽  
Saccharum Species ◽  
Subgroup Ii ◽  
Unrooted Phylogenetic Tree ◽  
Duplication Events

Sugarcane contributes ~80% of sugar production in the world and is an established biofuel crop. In working towards understanding the molecular basis of high sucrose accumulation, we have annotated and analysed the ATP-dependent phosphofructokinase (PFK) gene family that catalyses the phosphorylation of D-fructose 6-phosphate to D-fructose 1,6-bisphosphate. PFKs play an essential role in sucrose metabolism in plants and their expression patterns are unknown in sugarcane. In this study, based on the sorghum genome and sugarcane EST database, 10 PFK gene members were annotated and further verified by PCR using sugarcane genomic DNA. An unrooted phylogenetic tree was constructed with the deduced protein sequences of PFKs that were from the assembly of cDNA library of sugarcane and other plants. The results showed that gene duplication events and the retention rate after genome wide or segmental duplications occurred in higher frequency in monocots than in dicots and the genes in subgroup II of group III were likely originated from recent duplication events. Quantitative RT–PCR was performed to investigate the gene expression of 10 PFK genes in five tissues of three Saccharum species, including two developmental stages in leaves and three in culms. Of the PFK family members in sugarcane, ScPFK6, 7 and 8 appeared to be the primary isoforms based on the highly abundant expression of these three genes. ScPFK7 showed high expression level in the leaves, suggesting a potential role in sucrose metabolism. ScPFK8 had lower expression level in Saccharum officinarum L. than in the other two species, suggesting negative regulation of sucrose metabolism, which might have contributed to the high sugar content of S. officinarum. The genes in monocot specific subgroup II of group III, PFK7, 8 and 9, showed variation among the three Saccharum species, suggesting potential functional redundancy. Our results provide detailed annotation and analysis of the PFK gene family in sugarcane. Further elucidation of the role of ScPFK8 in the domestication process of sugarcane would be useful.

scholarly journals Developmental and functional studies of the SLC12 gene family members from Drosophila melanogaster

2010 ◽  
Vol 298 (1) ◽  
pp. C26-C37 ◽  
Author(s):  
Qifei Sun ◽  
E. Tian ◽  
R. James Turner ◽  
Kelly G. Ten Hagen
Keyword(s):  
Drosophila Melanogaster ◽  
Gene Family ◽  
Expression Patterns ◽  
Family Members ◽  
Specific Inhibitor ◽  
Insect Cell Line ◽  
Experimental Conditions ◽  
Functional Studies ◽  
Half Saturation Constant ◽  
Late Embryogenesis

The electroneutral cation-chloride cotransporter gene family, SLC12, contains nine members in vertebrates. These include seven sodium and/or potassium-coupled chloride transporters and two membrane proteins of unknown function. Although SLC12 family members have been identified in a number of lower species, the functional properties of these proteins are unknown. There are five SLC12 homologues in Drosophila melanogaster , including at least one member on each of the four main branches of the vertebrate phylogenetic tree. We have employed in situ hybridization to study the expression patterns of the Drosophila SLC12 proteins during embryonic development. Our studies indicate that all five members of this family are expressed during early embryogenesis ( stages 1–6), but that spatial and temporal expression patterns become more refined as development proceeds. Expression during late embryogenesis was seen predominantly in the ventral nerve cord, salivary gland, gut, and anal pad. In parallel studies, we have carried out transport assays on each of the five Drosophila homologues, expressed as recombinant proteins in the cultured insect cell line High Five. Under our experimental conditions, we found that only one of these proteins, CG4357, transported the potassium congener 86Rb. Additional experiments established that rubidium transport via CG4357 was saturable ( Km = 0.29 ± 0.05 mM), sodium-dependent (half-saturation constant = 53 ± 11 mM), chloride-dependent (half-saturation constant = 48 ± 5 mM), and potently inhibited by bumetanide (inhibitor constant = 1.17 ± 0.08 μM), a specific inhibitor of Na+-K+-2Cl− cotransporters. Taken together, our results provide strong evidence that CG4357 is an insect ortholog of the vertebrate Na+-K+-2Cl− cotransporters.

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