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Genome-wide identification, molecular cloning, expression profiling and posttranscriptional regulation analysis of the Argonaute gene family in Salvia miltiorrhiza, an emerging model medicinal plant.

Shao F, Lu S - BMC Genomics (2013)

Bottom Line: The results implied that some SmAGOs, such as SmAGO1, SmAGO2, SmAGO3, SmAGO7 and SmAGO10, probably played similar roles as their counterparts in Arabidopsis; whereas the others could be more species-specialized.Using the modified 5'-RACE method, we confirmed that SmAGO1 and SmAGO2 were targeted by S. miltiorrhiza miR168a/b and miR403, respectively.The results provide useful information for further elucidation of gene silencing pathways in S. miltiorrhiza.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences & Peking Union Medical College, No 151, Malianwa North Road, Haidian District, Beijing 100193, China.

ABSTRACT

Background: Argonaute (AGO) is the core component of RNA-induced silencing complex. The AGO gene family has been analyzed in various plant species; however, there is no report about AGOs in the well-known Traditional Chinese Medicine (TCM) plant, Salvia miltiorrhiza.

Results: Through a genome-wide analysis, we identified ten SmAGO genes in S. miltiorrhiza. Full-length cDNAs of all SmAGOs were subsequently cloned and sequenced. These SmAGOs were characterized using a comprehensive approach. Sequence features, gene structures and conserved domains were analyzed by the comparison of SmAGOs and AtAGOs. Phylogenetic relationships among AGO proteins from S. miltiorrhiza, Arabidopsis and rice were revealed. The expression levels of SmAGO genes in various tissues of S. miltiorrhiza were investigated. The results implied that some SmAGOs, such as SmAGO1, SmAGO2, SmAGO3, SmAGO7 and SmAGO10, probably played similar roles as their counterparts in Arabidopsis; whereas the others could be more species-specialized. It suggests the conservation and diversity of AGOs in plants. Additionally, we identified a total of 24 hairpin structures, representing six miRNA gene families, to be miRNA precursors. Using the modified 5'-RACE method, we confirmed that SmAGO1 and SmAGO2 were targeted by S. miltiorrhiza miR168a/b and miR403, respectively. It suggests the conservation of AGO1-miR168 and AGO2-miR403 regulatory modules in S. miltiorrhiza and Arabidopsis.

Conclusions: This is the first attempt to explore SmAGOs and miRNAs in S. miltiorrhiza. The results provide useful information for further elucidation of gene silencing pathways in S. miltiorrhiza.

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Expression of SmAGOs in flowers (Fl), leaves (Le), stems (St) and roots (Rt) of S. miltiorrhiza. Fold changes of SmAGO expression are shown. Expression levels were quantified by qRT-PCR. SmUBQ10 was used as a reference gene. The levels in roots were arbitrarily set to 1. Error bars represent the standard deviations of three technical PCR replicates.
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Figure 4: Expression of SmAGOs in flowers (Fl), leaves (Le), stems (St) and roots (Rt) of S. miltiorrhiza. Fold changes of SmAGO expression are shown. Expression levels were quantified by qRT-PCR. SmUBQ10 was used as a reference gene. The levels in roots were arbitrarily set to 1. Error bars represent the standard deviations of three technical PCR replicates.

Mentions: The expression of SmAGO genes in flowers, leaves, stems and roots of 2-year-old, field nursery-grown S. miltiorrhiza was analyzed using quantitative RT-PCR technology. SmUBQ10 was chosen as an endogenous control as previously described [26]. All of ten SmAGOs were expressed in S. miltiorrhiza tissues analyzed, whereas differential expression patterns were observed (Figure 4). SmAGO1 and SmAGO10 clustered in the AGO1 subgroup were expressed in flowers, leaves, stems and roots as their Arabidopsis counterparts, AtAGO1 and AtAGO10, respectively [11]. It is consistent with the ubiquitous roles of AGOs in the AGO1 subgroup. Although SmAGO2, SmAGO3 and SmAGO7 were clustered in the ZIPPY subgroup, their expression patterns were distinct (Figure 4). On the other hand, the expression patterns of SmAGO2, SmAGO3 and SmAGO7 were quite similar to their Arabidopsis counterparts, AtAGO3, AtAGO2, and AtAGO7, respectively [11]. By contrast, the expression pattern of SmAGO5 was distinct with their Arabidopsis and rice counterparts in the MEL1 subgroup. SmAGO5 showed the highest expression in roots, followed by stems, less in flowers and leaves. It is distinct with its counterparts, AtAGO5 in Arabidopsis and OsMEL1, OsAGO12, OsAGO13 and OsAGO14 in rice, showing specifically expression in reproductive tissues (Figure 4) [34]. Similar to the SmAGOs in the ZIPPY subgroup, SmAGO4, SmAGO6, SmAGO8 and SmAGO9 clustered in the AGO4 subgroup exhibited divergent expression patterns (Figure 4). SmAGO8 was more flower-specific, an expression pattern similar with SmAGO7 in the ZIPPY subgroup; whereas, SmAGO6 and SmAGO9 were more root-specific. The results indicate the functional conservation and diversity of AGOs.


Genome-wide identification, molecular cloning, expression profiling and posttranscriptional regulation analysis of the Argonaute gene family in Salvia miltiorrhiza, an emerging model medicinal plant.

Shao F, Lu S - BMC Genomics (2013)

Expression of SmAGOs in flowers (Fl), leaves (Le), stems (St) and roots (Rt) of S. miltiorrhiza. Fold changes of SmAGO expression are shown. Expression levels were quantified by qRT-PCR. SmUBQ10 was used as a reference gene. The levels in roots were arbitrarily set to 1. Error bars represent the standard deviations of three technical PCR replicates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750313&req=5

Figure 4: Expression of SmAGOs in flowers (Fl), leaves (Le), stems (St) and roots (Rt) of S. miltiorrhiza. Fold changes of SmAGO expression are shown. Expression levels were quantified by qRT-PCR. SmUBQ10 was used as a reference gene. The levels in roots were arbitrarily set to 1. Error bars represent the standard deviations of three technical PCR replicates.
Mentions: The expression of SmAGO genes in flowers, leaves, stems and roots of 2-year-old, field nursery-grown S. miltiorrhiza was analyzed using quantitative RT-PCR technology. SmUBQ10 was chosen as an endogenous control as previously described [26]. All of ten SmAGOs were expressed in S. miltiorrhiza tissues analyzed, whereas differential expression patterns were observed (Figure 4). SmAGO1 and SmAGO10 clustered in the AGO1 subgroup were expressed in flowers, leaves, stems and roots as their Arabidopsis counterparts, AtAGO1 and AtAGO10, respectively [11]. It is consistent with the ubiquitous roles of AGOs in the AGO1 subgroup. Although SmAGO2, SmAGO3 and SmAGO7 were clustered in the ZIPPY subgroup, their expression patterns were distinct (Figure 4). On the other hand, the expression patterns of SmAGO2, SmAGO3 and SmAGO7 were quite similar to their Arabidopsis counterparts, AtAGO3, AtAGO2, and AtAGO7, respectively [11]. By contrast, the expression pattern of SmAGO5 was distinct with their Arabidopsis and rice counterparts in the MEL1 subgroup. SmAGO5 showed the highest expression in roots, followed by stems, less in flowers and leaves. It is distinct with its counterparts, AtAGO5 in Arabidopsis and OsMEL1, OsAGO12, OsAGO13 and OsAGO14 in rice, showing specifically expression in reproductive tissues (Figure 4) [34]. Similar to the SmAGOs in the ZIPPY subgroup, SmAGO4, SmAGO6, SmAGO8 and SmAGO9 clustered in the AGO4 subgroup exhibited divergent expression patterns (Figure 4). SmAGO8 was more flower-specific, an expression pattern similar with SmAGO7 in the ZIPPY subgroup; whereas, SmAGO6 and SmAGO9 were more root-specific. The results indicate the functional conservation and diversity of AGOs.

Bottom Line: The results implied that some SmAGOs, such as SmAGO1, SmAGO2, SmAGO3, SmAGO7 and SmAGO10, probably played similar roles as their counterparts in Arabidopsis; whereas the others could be more species-specialized.Using the modified 5'-RACE method, we confirmed that SmAGO1 and SmAGO2 were targeted by S. miltiorrhiza miR168a/b and miR403, respectively.The results provide useful information for further elucidation of gene silencing pathways in S. miltiorrhiza.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences & Peking Union Medical College, No 151, Malianwa North Road, Haidian District, Beijing 100193, China.

ABSTRACT

Background: Argonaute (AGO) is the core component of RNA-induced silencing complex. The AGO gene family has been analyzed in various plant species; however, there is no report about AGOs in the well-known Traditional Chinese Medicine (TCM) plant, Salvia miltiorrhiza.

Results: Through a genome-wide analysis, we identified ten SmAGO genes in S. miltiorrhiza. Full-length cDNAs of all SmAGOs were subsequently cloned and sequenced. These SmAGOs were characterized using a comprehensive approach. Sequence features, gene structures and conserved domains were analyzed by the comparison of SmAGOs and AtAGOs. Phylogenetic relationships among AGO proteins from S. miltiorrhiza, Arabidopsis and rice were revealed. The expression levels of SmAGO genes in various tissues of S. miltiorrhiza were investigated. The results implied that some SmAGOs, such as SmAGO1, SmAGO2, SmAGO3, SmAGO7 and SmAGO10, probably played similar roles as their counterparts in Arabidopsis; whereas the others could be more species-specialized. It suggests the conservation and diversity of AGOs in plants. Additionally, we identified a total of 24 hairpin structures, representing six miRNA gene families, to be miRNA precursors. Using the modified 5'-RACE method, we confirmed that SmAGO1 and SmAGO2 were targeted by S. miltiorrhiza miR168a/b and miR403, respectively. It suggests the conservation of AGO1-miR168 and AGO2-miR403 regulatory modules in S. miltiorrhiza and Arabidopsis.

Conclusions: This is the first attempt to explore SmAGOs and miRNAs in S. miltiorrhiza. The results provide useful information for further elucidation of gene silencing pathways in S. miltiorrhiza.

Show MeSH