Identification and characterization of long non-coding RNA (lncRNA) in the developing seeds of Jatropha curcas

Abstract

Long non-coding RNAs (lncRNAs) play critical roles in plant development. However, the information of lncRNAs in Jatropha curcas remains largely unexplored. Thus, an attempt has been made in J. curcas to identify 1,850 lncRNAs based on deep sequencing of developing seeds at three typical stages. About ten percent lncRNAs (196 lncRNAs) were differentially expressed lncRNAs during seed developing process. Together with reverse transcription quantitative real-time PCR, the lncRNA expression analyses revealed the stage-specific expression patterns of some novel lncRNAs in J. curcas. The target genes of lncRNAs were annotated for their roles in various biological processes such as gene expression, metabolism, and cell growth. Besides, 10 lncRNAs were identified as the precursors of microRNAs and 26 lncRNAs were predicted to be the targets of Jatropha miRNAs. A total of 31 key lncRNAs play critical roles in the seed developing process in the context of cell growth and development, lipid metabolism, and seed maturation. Our study provides the first systematic study of lncRNAs in the developing seeds of J. curcas and facilitates the functional research of plant lncRNAs and the regulation of seed development.

Figure 1

The Venn diagram of the noncoding transcripts identified by CPC, CNCI, CPAT, and Pfam analyses. Each circle represents an analysis method for predicting lncRNA. The number in the circle represents the number of transcripts predicted to be lncRNAs. The intersection of four circles is taken as the final result of lncRNAs.

Figure 2

The types of lncRNA. The abscissa is the 4 types of lncRNA and the ordinate is the number of the corresponding lncRNA. LincRNA represents intergenic lncRNA.

Figure 3

The differentially expressed lncRNAs during seed development of J. curcas. Differential expression analyses of lncRNAs were performed based on FPKMs of lncRNAs in each sample, which were calculated using the length of lncRNAs (kilo-base) and mapped fragments (million). The variation of lncRNA levels in three different developmental stages, i.e., small (S), middle (M) and large (L) seeds were shown as the base-2 logarithm of ratios (log₂ fold change) between different developmental stages (L/M, M/S and L/S). The lncRNAs shown were differentially expressed lncRNAs because their expression levels between any two of the three different developmental stages changed significantly (|log₂ fold change|> 1 and p < 0.01).

Figure 4

The function category of KOG annotation for the targets of differentially expressed lncRNAs during seed development of J. curcas. X-axis represents KOG category. Y-axis indicates the number of lncRNA target genes. Gene function was annotated on the basis of KOG/COG (Clusters of Orthologous Groups of proteins). The genes with unknown function were not counted in. Upper panel, target genes of up-regulated lncRNAs; lower panel, target genes of down-regulated lncRNAs.

Figure 5

Key lncRNAs in the context of cell growth and development, lipid metabolism, and seed maturation of Jatropha developing seeds. Numbers before the arrows are lncRNA ID, and the targets of lncRNA are shown after the arrows. The lncRNAs might act as endogenous target mimics of miRNA, and the miRNA targets, are shown in red (For detail, see Supplementary Table S7).