Genome-Wide Identification and Characterization of UTR-Introns of Citrus sinensis

Abstract

Introns exist not only in coding sequences (CDSs) but also in untranslated regions (UTRs) of a gene. Recent studies in animals and model plants such as Arabidopsis have revealed that the UTR-introns (UIs) are widely presented in most genomes and involved in regulation of gene expression or RNA stability. In the present study, we identified introns at both 5′UTRs (5UIs) and 3′UTRs (3UIs) of sweet orange genes, investigated their size and nucleotide distribution characteristics, and explored the distribution of cis-elements in the UI sequences. Functional category of genes with predicted UIs were further analyzed using GO, KEGG, and PageMan enrichment. In addition, the organ-dependent splicing and abundance of selected UI-containing genes in root, leaf, and stem were experimentally determined. Totally, we identified 825 UI- and 570 3UI-containing transcripts, corresponding to 617 and 469 genes, respectively. Among them, 74 genes contain both 5UI and 3UI. Nucleotide distribution analysis showed that 5UI distribution is biased at both ends of 5′UTR whiles 3UI distribution is biased close to the start site of 3′UTR. Cis- elements analysis revealed that 5UI and 3UI sequences were rich of promoter-enhancing related elements, indicating that they might function in regulating the expression through them. Function enrichment analysis revealed that genes containing 5UI are significantly enriched in the RNA transport pathway. While, genes containing 3UI are significantly enriched in splicesome. Notably, many pentatricopeptide repeat-containing protein genes and the disease resistance genes were identified to be 3UI-containing. RT-PCR result confirmed the existence of UIs in the eight selected gene transcripts whereas alternative splicing events were found in some of them. Meanwhile, qRT-PCR result showed that UIs were differentially expressed among organs, and significant correlation was found between some genes and their UIs, for example: The expression of VPS28 and its 3UI was significantly negative correlated. This is the first report about the UIs in sweet orange from genome-wide level, which could provide evidence for further understanding of the role of UIs in gene expression regulation.

Genome-Scale Computational Identification and Characterization of UTR Introns in Atalantia buxifolia

“…However, the relative frequencies of short introns < 50 nucleotides and introns > 300 nucleotides of 3UIs and 5UIs were higher than CIs. Similar to sweet orange [42], the A. buxifolia 5UIs and 3UIs are more preferentially located at the stop ends of 5 UTRs and at the beginning of 3 UTRs, respectively.…”

“…Based on the annotation information of the A. buxifolia genome, introns in CDSs, 5 UTR and 3 UTR were separately extracted. Then, the 5UIs and 3UIs were identified from the A. buxifolia genome according to the method described by Shi et al [42]. Introns between UTR exons were extracted according to the genome annotation file, and introns showed retention in the exon region of any transcripts were excluded to ensure the identified UIs strictly exist in the intron regions.…”

“…Information of the identified 5UIs and 3UIs were shown in Additional file Tables S1 and S2, respectively. UI density, position preference, length and nucleotide composition statistical analysis were performed using Perl, and figures were drawn using ggplot2 [42].…”

“…To show the nucleotide bias around the donor and acceptor sites of 5UIs, CIs and 3UIs, the sequence logos were used [49]. Results showed that both the A. buxifolia UIs and CIs possess A/T-rich element around both donor and receptor sites [42]. Moreover, GT-AG was found to be the major splice site pair in both A. buxifolia 5 UTRs (98.32%) and 3 UTRs (98.26%), followed by GC-AG splice site pair (accounting for 1.67% of 5 UTRs and 1.73% of 3 UTRs) (Figure 2).…”

“…Cenik et al [41] discovered that human genes with regulatory roles were surprisingly rich in 5UI. Shi et al [42] analyzed the UIs characteristics and revealed UIs' differential expression in different organs of sweet orange. These findings can provide information for evolution and functional significances of UIs.…”