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Small RNA and Degradome Sequencing Reveal Complex Roles of miRNAs and Their Targets in Developing Wheat Grains.

Li T, Ma L, Geng Y, Hao C, Chen X, Zhang X - PLoS ONE (2015)

Bottom Line: A comparison of the miRNAomes revealed that 55 miRNA families were differentially expressed during the grain development.Predicted and validated targets of these development-related miRNAs are involved in different cellular responses and metabolic processes including cell proliferation, auxin signaling, nutrient metabolism and gene expression.This study provides insight into the complex roles of miRNAs and their targets in regulating wheat grain development.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Crop Gene Resources and Germplasm Enhancement, Ministry of Agriculture / Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, China.

ABSTRACT
Plant microRNAs (miRNAs) have been shown to play critical roles in plant development. In this study, we employed small RNA combined with degradome sequencing to survey development-related miRNAs and their validated targets during wheat grain development. A total of 186 known miRNAs and 37 novel miRNAs were identified in four small RNA libraries. Moreover, a miRNA-like long hairpin locus was first identified to produce 21~22-nt phased siRNAs that act in trans to cleave target mRNAs. A comparison of the miRNAomes revealed that 55 miRNA families were differentially expressed during the grain development. Predicted and validated targets of these development-related miRNAs are involved in different cellular responses and metabolic processes including cell proliferation, auxin signaling, nutrient metabolism and gene expression. This study provides insight into the complex roles of miRNAs and their targets in regulating wheat grain development.

No MeSH data available.


Verified targets of conserved development-related miRNAs.(A) Confirmation of target mRNA cleavage by degradome sequencing. The t-plots show sequence abundances in the position of the target transcripts, with red lines indicating sequence reads consistent with miRNA-directed cleavage. All the data of the t-plots originated from degradome library II. (B) Mapping of target mRNA cleavage sites using 5′-RACE. Arrows indicate cleavage sites and the number shows the frequency of sequenced clones.
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pone.0139658.g004: Verified targets of conserved development-related miRNAs.(A) Confirmation of target mRNA cleavage by degradome sequencing. The t-plots show sequence abundances in the position of the target transcripts, with red lines indicating sequence reads consistent with miRNA-directed cleavage. All the data of the t-plots originated from degradome library II. (B) Mapping of target mRNA cleavage sites using 5′-RACE. Arrows indicate cleavage sites and the number shows the frequency of sequenced clones.

Mentions: To further investigate the functions of the 55 development-related miRNAs differentially expressed during grain development, we performed target prediction combined with degradome analysis (S6 Table). The targets for highly conserved miRNAs were mostly supported by degradome sequencing data (S6 Table). Most verified targets/miRNA modules were highly conserved among monocots and dicots, such as SPLs/miR156, NACs/miR164, HOXs/miR166, ARFs/miR167, SCL1/miR171, TCPs/miR319, GRFs/miR396, SPX/miR827 (Fig 4A). In particular, miR164 was validated to regulate a new class of targets encoding phytosulfokine-alpha 1 precursor (PSK1). This regulation is also present in rice but absent in Arabidopsis [40], indicating it is less conserved and emerged more recently in evolution. To further confirm the degradome data, we selected three targets of conserved miRNAs for validation using rapid amplification of 5′ cDNA ends (5′ RACE). As shown in Fig 4B, the major cleavage sites of TC398770 (SCL1) and TC398226 (TCP) were present at the canonical position, consistent with corresponding degradome data (Fig 4A). In addition, miR156-mediated cleavage of Ta.6374 (SPL13) mRNA, which was weakly identified by degradome data (Fig 4A), was also verified by 5′ RACE, indicating that these targets of conserved miRNAs can be well supported by degradome data combined with 5′ RACE.


Small RNA and Degradome Sequencing Reveal Complex Roles of miRNAs and Their Targets in Developing Wheat Grains.

Li T, Ma L, Geng Y, Hao C, Chen X, Zhang X - PLoS ONE (2015)

Verified targets of conserved development-related miRNAs.(A) Confirmation of target mRNA cleavage by degradome sequencing. The t-plots show sequence abundances in the position of the target transcripts, with red lines indicating sequence reads consistent with miRNA-directed cleavage. All the data of the t-plots originated from degradome library II. (B) Mapping of target mRNA cleavage sites using 5′-RACE. Arrows indicate cleavage sites and the number shows the frequency of sequenced clones.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591353&req=5

pone.0139658.g004: Verified targets of conserved development-related miRNAs.(A) Confirmation of target mRNA cleavage by degradome sequencing. The t-plots show sequence abundances in the position of the target transcripts, with red lines indicating sequence reads consistent with miRNA-directed cleavage. All the data of the t-plots originated from degradome library II. (B) Mapping of target mRNA cleavage sites using 5′-RACE. Arrows indicate cleavage sites and the number shows the frequency of sequenced clones.
Mentions: To further investigate the functions of the 55 development-related miRNAs differentially expressed during grain development, we performed target prediction combined with degradome analysis (S6 Table). The targets for highly conserved miRNAs were mostly supported by degradome sequencing data (S6 Table). Most verified targets/miRNA modules were highly conserved among monocots and dicots, such as SPLs/miR156, NACs/miR164, HOXs/miR166, ARFs/miR167, SCL1/miR171, TCPs/miR319, GRFs/miR396, SPX/miR827 (Fig 4A). In particular, miR164 was validated to regulate a new class of targets encoding phytosulfokine-alpha 1 precursor (PSK1). This regulation is also present in rice but absent in Arabidopsis [40], indicating it is less conserved and emerged more recently in evolution. To further confirm the degradome data, we selected three targets of conserved miRNAs for validation using rapid amplification of 5′ cDNA ends (5′ RACE). As shown in Fig 4B, the major cleavage sites of TC398770 (SCL1) and TC398226 (TCP) were present at the canonical position, consistent with corresponding degradome data (Fig 4A). In addition, miR156-mediated cleavage of Ta.6374 (SPL13) mRNA, which was weakly identified by degradome data (Fig 4A), was also verified by 5′ RACE, indicating that these targets of conserved miRNAs can be well supported by degradome data combined with 5′ RACE.

Bottom Line: A comparison of the miRNAomes revealed that 55 miRNA families were differentially expressed during the grain development.Predicted and validated targets of these development-related miRNAs are involved in different cellular responses and metabolic processes including cell proliferation, auxin signaling, nutrient metabolism and gene expression.This study provides insight into the complex roles of miRNAs and their targets in regulating wheat grain development.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Crop Gene Resources and Germplasm Enhancement, Ministry of Agriculture / Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, China.

ABSTRACT
Plant microRNAs (miRNAs) have been shown to play critical roles in plant development. In this study, we employed small RNA combined with degradome sequencing to survey development-related miRNAs and their validated targets during wheat grain development. A total of 186 known miRNAs and 37 novel miRNAs were identified in four small RNA libraries. Moreover, a miRNA-like long hairpin locus was first identified to produce 21~22-nt phased siRNAs that act in trans to cleave target mRNAs. A comparison of the miRNAomes revealed that 55 miRNA families were differentially expressed during the grain development. Predicted and validated targets of these development-related miRNAs are involved in different cellular responses and metabolic processes including cell proliferation, auxin signaling, nutrient metabolism and gene expression. This study provides insight into the complex roles of miRNAs and their targets in regulating wheat grain development.

No MeSH data available.