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Computational Identification and Systematic Classification of Novel Cytochrome P450 Genes in Salvia miltiorrhiza.

Chen H, Wu B, Nelson DR, Wu K, Liu C - PLoS ONE (2014)

Bottom Line: The RNA-Seq results showed that 35 CYP450 genes were co-expressed with CYP76AH1, a marker gene for tanshinone biosynthesis, using r≥0.9 as a cutoff.Comparing against the KEGG database, 10 CYP450 genes were found to be associated with diterpenoid biosynthesis.Moreover, we found that 15 CYP450 genes were possibly regulated by antisense transcripts (r≥0.9 or r≤-0.9).

View Article: PubMed Central - PubMed

Affiliation: Center for Bioinformatics, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.

ABSTRACT
Salvia miltiorrhiza is one of the most economically important medicinal plants. Cytochrome P450 (CYP450) genes have been implicated in the biosynthesis of its active components. However, only a dozen full-length CYP450 genes have been described, and there is no systematic classification of CYP450 genes in S. miltiorrhiza. We obtained 77,549 unigenes from three tissue types of S. miltiorrhiza using RNA-Seq technology. Combining our data with previously identified CYP450 sequences and scanning with the CYP450 model from Pfam resulted in the identification of 116 full-length and 135 partial-length CYP450 genes. The 116 genes were classified into 9 clans and 38 families using standard criteria. The RNA-Seq results showed that 35 CYP450 genes were co-expressed with CYP76AH1, a marker gene for tanshinone biosynthesis, using r≥0.9 as a cutoff. The expression profiles for 16 of 19 randomly selected CYP450 obtained from RNA-Seq were validated by qRT-PCR. Comparing against the KEGG database, 10 CYP450 genes were found to be associated with diterpenoid biosynthesis. Considering all the evidence, 3 CYP450 genes were identified to be potentially involved in terpenoid biosynthesis. Moreover, we found that 15 CYP450 genes were possibly regulated by antisense transcripts (r≥0.9 or r≤-0.9). Lastly, a web resource (SMCYP450, http://www.herbalgenomics.org/samicyp450) was set up, which allows users to browse, search, retrieve and compare CYP450 genes and can serve as a centralized resource.

No MeSH data available.


Phylogenetic analyses of non-A-type CYP450s from S. lycopersicum and S. miltiorrhiza.The Neighbor-Joining tree was constructed using the MEGA6 package. The spokes corresponding to Sm and Sl CYP450s are shown in green and black, respectively. The shaded genes are described in the text. Sm: S. miltiorrhiza and Sl: S. lycopersicum.
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pone-0115149-g004: Phylogenetic analyses of non-A-type CYP450s from S. lycopersicum and S. miltiorrhiza.The Neighbor-Joining tree was constructed using the MEGA6 package. The spokes corresponding to Sm and Sl CYP450s are shown in green and black, respectively. The shaded genes are described in the text. Sm: S. miltiorrhiza and Sl: S. lycopersicum.

Mentions: As described above, members from several families, such as CYP92A, CYP736, CYP749, CYP728 and CYP727, were missing in A. thaliana, suggesting that A. thaliana is not the best reference for S. miltiorrhiza in comparative studies of CYP450 genes. [29]. Instead, the Solanum lycopersicum CYPome was chosen for comparison because S. lycopersicum belongs to Solanales, the order closest to Lamiales, which includes the family lamiaceae and has well-annotated CYP450 genes. The sequences of 271 S. lycopersicum CYP450 proteins and the 116 full-length S. miltiorrhiza CYP450 proteins were used to construct Neighbor-Joining (NJ) trees for A-type (Fig. 3) and non- A-type (Fig. 4) CYP450s, respectively, using the MEGA6 program. The trees showed that among the 116 full-length CYP450 genes, 65 (56.0%) were A-type and distributed into 17 families. The remaining 51 (44.0%) full-length CYP450s were non-A-type, belonging to 21 families and 9 clans. Genes belonging to the CYP71 clan have been found to be involved in the biosynthesis of secondary metabolites or natural products [15], [33]. It has been reported that CYP76AH1 gene is involved in tanshinone biosynthesis [11]. The large number of subfamilies found in the CYP76 family indicates a diversity of functions, in which even genes within the same subfamily may have unique functions. The phylogenetic tree for non-A-type CYP450s is shown in Fig. 4. These include a much more divergent group of sequences covering the 8 remaining clans, which function in sterol, carotenoid, oxylipin, fatty acid and hormone metabolism [34] as well as secondary metabolite synthesis. It should be noted that most genes belonging to family CYP76 are clustered on different branches for S. miltiorrhiza and S. lycopersicum (Fig. 3, shaded area), suggesting that this family underwent independent expansion after the divergence of Lamiales and Solanales, possibly to meet unique functional needs.


Computational Identification and Systematic Classification of Novel Cytochrome P450 Genes in Salvia miltiorrhiza.

Chen H, Wu B, Nelson DR, Wu K, Liu C - PLoS ONE (2014)

Phylogenetic analyses of non-A-type CYP450s from S. lycopersicum and S. miltiorrhiza.The Neighbor-Joining tree was constructed using the MEGA6 package. The spokes corresponding to Sm and Sl CYP450s are shown in green and black, respectively. The shaded genes are described in the text. Sm: S. miltiorrhiza and Sl: S. lycopersicum.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4262458&req=5

pone-0115149-g004: Phylogenetic analyses of non-A-type CYP450s from S. lycopersicum and S. miltiorrhiza.The Neighbor-Joining tree was constructed using the MEGA6 package. The spokes corresponding to Sm and Sl CYP450s are shown in green and black, respectively. The shaded genes are described in the text. Sm: S. miltiorrhiza and Sl: S. lycopersicum.
Mentions: As described above, members from several families, such as CYP92A, CYP736, CYP749, CYP728 and CYP727, were missing in A. thaliana, suggesting that A. thaliana is not the best reference for S. miltiorrhiza in comparative studies of CYP450 genes. [29]. Instead, the Solanum lycopersicum CYPome was chosen for comparison because S. lycopersicum belongs to Solanales, the order closest to Lamiales, which includes the family lamiaceae and has well-annotated CYP450 genes. The sequences of 271 S. lycopersicum CYP450 proteins and the 116 full-length S. miltiorrhiza CYP450 proteins were used to construct Neighbor-Joining (NJ) trees for A-type (Fig. 3) and non- A-type (Fig. 4) CYP450s, respectively, using the MEGA6 program. The trees showed that among the 116 full-length CYP450 genes, 65 (56.0%) were A-type and distributed into 17 families. The remaining 51 (44.0%) full-length CYP450s were non-A-type, belonging to 21 families and 9 clans. Genes belonging to the CYP71 clan have been found to be involved in the biosynthesis of secondary metabolites or natural products [15], [33]. It has been reported that CYP76AH1 gene is involved in tanshinone biosynthesis [11]. The large number of subfamilies found in the CYP76 family indicates a diversity of functions, in which even genes within the same subfamily may have unique functions. The phylogenetic tree for non-A-type CYP450s is shown in Fig. 4. These include a much more divergent group of sequences covering the 8 remaining clans, which function in sterol, carotenoid, oxylipin, fatty acid and hormone metabolism [34] as well as secondary metabolite synthesis. It should be noted that most genes belonging to family CYP76 are clustered on different branches for S. miltiorrhiza and S. lycopersicum (Fig. 3, shaded area), suggesting that this family underwent independent expansion after the divergence of Lamiales and Solanales, possibly to meet unique functional needs.

Bottom Line: The RNA-Seq results showed that 35 CYP450 genes were co-expressed with CYP76AH1, a marker gene for tanshinone biosynthesis, using r≥0.9 as a cutoff.Comparing against the KEGG database, 10 CYP450 genes were found to be associated with diterpenoid biosynthesis.Moreover, we found that 15 CYP450 genes were possibly regulated by antisense transcripts (r≥0.9 or r≤-0.9).

View Article: PubMed Central - PubMed

Affiliation: Center for Bioinformatics, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.

ABSTRACT
Salvia miltiorrhiza is one of the most economically important medicinal plants. Cytochrome P450 (CYP450) genes have been implicated in the biosynthesis of its active components. However, only a dozen full-length CYP450 genes have been described, and there is no systematic classification of CYP450 genes in S. miltiorrhiza. We obtained 77,549 unigenes from three tissue types of S. miltiorrhiza using RNA-Seq technology. Combining our data with previously identified CYP450 sequences and scanning with the CYP450 model from Pfam resulted in the identification of 116 full-length and 135 partial-length CYP450 genes. The 116 genes were classified into 9 clans and 38 families using standard criteria. The RNA-Seq results showed that 35 CYP450 genes were co-expressed with CYP76AH1, a marker gene for tanshinone biosynthesis, using r≥0.9 as a cutoff. The expression profiles for 16 of 19 randomly selected CYP450 obtained from RNA-Seq were validated by qRT-PCR. Comparing against the KEGG database, 10 CYP450 genes were found to be associated with diterpenoid biosynthesis. Considering all the evidence, 3 CYP450 genes were identified to be potentially involved in terpenoid biosynthesis. Moreover, we found that 15 CYP450 genes were possibly regulated by antisense transcripts (r≥0.9 or r≤-0.9). Lastly, a web resource (SMCYP450, http://www.herbalgenomics.org/samicyp450) was set up, which allows users to browse, search, retrieve and compare CYP450 genes and can serve as a centralized resource.

No MeSH data available.