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. 2006 May 19:7:120.
doi: 10.1186/1471-2164-7-120.

Effect of 5'UTR introns on gene expression in Arabidopsis thaliana

Betty Y W Chung  1 , Cas Simons, Andrew E Firth, Chris M Brown, Roger P Hellens
Affiliations

Effect of 5'UTR introns on gene expression in Arabidopsis thaliana

Betty Y W Chung et al. BMC Genomics. .

Abstract

Background: The majority of introns in gene transcripts are found within the coding sequences (CDSs). A small but significant fraction of introns are also found to reside within the untranslated regions (5'UTRs and 3'UTRs) of expressed sequences. Alignment of the whole genome and expressed sequence tags (ESTs) of the model plant Arabidopsis thaliana has identified introns residing in both coding and non-coding regions of the genome.

Results: A bioinformatic analysis revealed some interesting observations: (1) the density of introns in 5'UTRs is similar to that in CDSs but much higher than that in 3'UTRs; (2) the 5'UTR introns are preferentially located close to the initiating ATG codon; (3) introns in the 5'UTRs are, on average, longer than introns in the CDSs and 3'UTRs; and (4) 5'UTR introns have a different nucleotide composition to that of CDS and 3'UTR introns. Furthermore, we show that the 5'UTR intron of the A. thaliana EF1alpha-A3 gene affects the gene expression and the size of the 5'UTR intron influences the level of gene expression.

Conclusion: Introns within the 5'UTR show specific features that distinguish them from introns that reside within the coding sequence and the 3'UTR. In the EF1alpha-A3 gene, the presence of a long intron in the 5'UTR is sufficient to enhance gene expression in plants in a size dependent manner.

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Figures

Figure 1
Figure 1
Comparison of the length distributions of 5'UTR,3'UTR and CDS introns. Bar graph comparing the length distribution of 5'UTR (grey), CDS (black) and 3'UTR (white) introns. The x-axis labels give the mid-points of the length range that each bar covers (e.g. if the range is 51–100 nucleotides, the mid-point is 75.5, bin size = 50 nucleotides).
Figure 2
Figure 2
Distribution of 5'UTR intron positions relative to the start and end points of the associated UTRs. Pink bars represent the observed positions of 5'UTR introns relative to the end of the 5'UTR (i.e. the start codon ATG). Black bars represent the observed positions of 5'UTR introns relative to the beginning of the 5'UTR. The line represents the random model expected values (identical for both the black and pink bars). The positions are the intron excision points in the spliced transcript. The x-axis shows the mid-points of the length range that each bar covers (e.g. if the range is 41–60 nucleotides, the mid-point is 50.5, bin size = 20 nucleotides).
Figure 3
Figure 3
Sequence logos showing the nucleotide bias around the donor site of 5'UTR, CDS and 3'UTR introns. The x-axis refers to bases from the beginning of the intron, letter heights reflect the nucleotide bias at each position. Only 5 nucleotides of exon and 25 nucleotides of intron sequence at the donor site were included in the sequence logo as deviation of the nucleotide usage from background levels is not apparent outside these regions.
Figure 4
Figure 4
Sequence logos showing the nucleotide bias around the acceptor site of 5'UTR, CDS and 3'UTR introns. The x-axis refers to bases from the beginning of the intron, letter heights reflect the nucleotide bias at each position. Only 2 nucleotides of exon and 25 nucleotides of intron sequence at the acceptor site were included in the sequence logo as deviation of the nucleotide usage from background levels is not apparent outside these regions.
Figure 5
Figure 5
Schematic diagram of the plasmids, namely pGEF1I and pGEF1Idel. The plasmidswere used in the transient assays shown in Figure 6.
Figure 6
Figure 6
Scatter plot of the actual firefly and Renilla counts. Transient effect of 5'UTR intron (602 nucleotides) presence on the gene expression. The relative expression (± standard errors) is 0.0340 ± 0.0016 with the intron (triangles, pGEF1I) and 0.0139 ± 0.0008 without the intron (squares, pGEF1Idel).
Figure 7
Figure 7
Absolute firefly luciferase activity of transgenic plants. Average of absolute firefly luciferase activity for two T1 leaves from each of 10 independent plants transformed with pGEF1Ikan, and 7 independent plants transformed with pGEF1Idelkan. Error bars are standard deviation of two T1 leaves from the same plant.
Figure 8
Figure 8
Schematic diagram of AtEF1α-A3 5'UTR intron deletion series. A series of truncated EF1α-A3 5'UTR introns from AT1G07940 were generated. Fifty nucleotides up and downstream of the intron acceptor and donor sites were maintained for splicing efficiency. The deletions are grouped according to the position of the deletion point relative to the 3' region of the intron. Normalised relative luciferase values represent the regression of six independent measurements (see also Figure 9).
Figure 9
Figure 9
Luciferase expression of AtEF1α-A3 5'UTR intron deletions. On the x-axis, the number below each bar indicates the plasmid tested (Figure 8). Each bar represents the regression from six independent LUC-REN expression measurements. Error bars are standard errors.
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