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Cross-species EST alignments reveal novel and conserved alternative splicing events in legumes.

Wang BB, O'Toole M, Brendel V, Young ND - BMC Plant Biol. (2008)

Bottom Line: Intron retention is the most common form of AS in all four plant species (~50%), with slightly lower frequency in legumes compared to Arabidopsis and rice.The results also indicate that the frequency of AS in plants is comparable to that observed in mammals.Finally, our results highlight the importance of normalizing EST levels when estimating the frequency of alternative splicing.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Plant Pathology, University of Minnesota, St, Paul, MN 55108, USA. wangx741@umn.edu

ABSTRACT

Background: Although originally thought to be less frequent in plants than in animals, alternative splicing (AS) is now known to be widespread in plants. Here we report the characteristics of AS in legumes, one of the largest and most important plant families, based on EST alignments to the genome sequences of Medicago truncatula (Mt) and Lotus japonicus (Lj).

Results: Based on cognate EST alignments alone, the observed frequency of alternatively spliced genes is lower in Mt (approximately 10%, 1,107 genes) and Lj (approximately 3%, 92 genes) than in Arabidopsis and rice (both around 20%). However, AS frequencies are comparable in all four species if EST levels are normalized. Intron retention is the most common form of AS in all four plant species (~50%), with slightly lower frequency in legumes compared to Arabidopsis and rice. This differs notably from vertebrates, where exon skipping is most common. To uncover additional AS events, we aligned ESTs from other legume species against the Mt genome sequence. In this way, 248 additional Mt genes were predicted to be alternatively spliced. We also identified 22 AS events completely conserved in two or more plant species.

Conclusion: This study extends the range of plant taxa shown to have high levels of AS, confirms the importance of intron retention in plants, and demonstrates the utility of using ESTs from related species in order to identify novel and conserved AS events. The results also indicate that the frequency of AS in plants is comparable to that observed in mammals. Finally, our results highlight the importance of normalizing EST levels when estimating the frequency of alternative splicing.

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Correlation between AS frequency and EST coverage. The x-axis indicates groups of genes with certain numbers of ESTs. The primary y-axis for the bar graph indicates total number of genes within each group. The secondary y-axis for the line graph indicates the fraction of alternatively spliced genes for the group. Note that different bin sizes were used to keep the number of genes in each group greater than 500 in At and Os. AS data from groups with fewer than 80 genes in Mt were removed to reduce noise. Lj data were not shown as only the first six groups have more than 80 genes.
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Figure 2: Correlation between AS frequency and EST coverage. The x-axis indicates groups of genes with certain numbers of ESTs. The primary y-axis for the bar graph indicates total number of genes within each group. The secondary y-axis for the line graph indicates the fraction of alternatively spliced genes for the group. Note that different bin sizes were used to keep the number of genes in each group greater than 500 in At and Os. AS data from groups with fewer than 80 genes in Mt were removed to reduce noise. Lj data were not shown as only the first six groups have more than 80 genes.

Mentions: To compare the frequency of alternative splicing between different species, earlier studies relied on 10 randomly selected ESTs per gene as a basis for estimating AS frequency [4]. Here, only a small fraction (10–20%) of legume genes were covered by 10 or more ESTs, so this approach was not practical. Instead, we plotted the AS frequency for all groups of genes with similar EST coverage in different species, as shown in Figure 2. Mt categories with fewer than 80 genes total were removed to reduce noise due to small sample size, and Lj data are not included at all, as sample size was uniformly too small. When analyzed in this way, the fractions of alternatively spliced genes are similar regardless of species for nearly all size classes. For genes with four ESTs (the median EST number per gene in Mt), the observed AS frequency is 6–12% in Mt, At, and Os alike. For genes with nine to 11 ESTs (the median EST number per gene in Os and At), 15–23% are alternatively spliced. In general, the fraction of alternatively spliced genes keeps increasing with increasing transcript coverage, eventually reaching 66% in Os and 46% in At for genes with hundreds of ESTs, a levels similar to those observed in mammals [38,39]. Interestingly, the AS level in Os is consistently over 10% higher than in At in genes with more than 40 supporting ESTs.


Cross-species EST alignments reveal novel and conserved alternative splicing events in legumes.

Wang BB, O'Toole M, Brendel V, Young ND - BMC Plant Biol. (2008)

Correlation between AS frequency and EST coverage. The x-axis indicates groups of genes with certain numbers of ESTs. The primary y-axis for the bar graph indicates total number of genes within each group. The secondary y-axis for the line graph indicates the fraction of alternatively spliced genes for the group. Note that different bin sizes were used to keep the number of genes in each group greater than 500 in At and Os. AS data from groups with fewer than 80 genes in Mt were removed to reduce noise. Lj data were not shown as only the first six groups have more than 80 genes.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2277414&req=5

Figure 2: Correlation between AS frequency and EST coverage. The x-axis indicates groups of genes with certain numbers of ESTs. The primary y-axis for the bar graph indicates total number of genes within each group. The secondary y-axis for the line graph indicates the fraction of alternatively spliced genes for the group. Note that different bin sizes were used to keep the number of genes in each group greater than 500 in At and Os. AS data from groups with fewer than 80 genes in Mt were removed to reduce noise. Lj data were not shown as only the first six groups have more than 80 genes.
Mentions: To compare the frequency of alternative splicing between different species, earlier studies relied on 10 randomly selected ESTs per gene as a basis for estimating AS frequency [4]. Here, only a small fraction (10–20%) of legume genes were covered by 10 or more ESTs, so this approach was not practical. Instead, we plotted the AS frequency for all groups of genes with similar EST coverage in different species, as shown in Figure 2. Mt categories with fewer than 80 genes total were removed to reduce noise due to small sample size, and Lj data are not included at all, as sample size was uniformly too small. When analyzed in this way, the fractions of alternatively spliced genes are similar regardless of species for nearly all size classes. For genes with four ESTs (the median EST number per gene in Mt), the observed AS frequency is 6–12% in Mt, At, and Os alike. For genes with nine to 11 ESTs (the median EST number per gene in Os and At), 15–23% are alternatively spliced. In general, the fraction of alternatively spliced genes keeps increasing with increasing transcript coverage, eventually reaching 66% in Os and 46% in At for genes with hundreds of ESTs, a levels similar to those observed in mammals [38,39]. Interestingly, the AS level in Os is consistently over 10% higher than in At in genes with more than 40 supporting ESTs.

Bottom Line: Intron retention is the most common form of AS in all four plant species (~50%), with slightly lower frequency in legumes compared to Arabidopsis and rice.The results also indicate that the frequency of AS in plants is comparable to that observed in mammals.Finally, our results highlight the importance of normalizing EST levels when estimating the frequency of alternative splicing.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Plant Pathology, University of Minnesota, St, Paul, MN 55108, USA. wangx741@umn.edu

ABSTRACT

Background: Although originally thought to be less frequent in plants than in animals, alternative splicing (AS) is now known to be widespread in plants. Here we report the characteristics of AS in legumes, one of the largest and most important plant families, based on EST alignments to the genome sequences of Medicago truncatula (Mt) and Lotus japonicus (Lj).

Results: Based on cognate EST alignments alone, the observed frequency of alternatively spliced genes is lower in Mt (approximately 10%, 1,107 genes) and Lj (approximately 3%, 92 genes) than in Arabidopsis and rice (both around 20%). However, AS frequencies are comparable in all four species if EST levels are normalized. Intron retention is the most common form of AS in all four plant species (~50%), with slightly lower frequency in legumes compared to Arabidopsis and rice. This differs notably from vertebrates, where exon skipping is most common. To uncover additional AS events, we aligned ESTs from other legume species against the Mt genome sequence. In this way, 248 additional Mt genes were predicted to be alternatively spliced. We also identified 22 AS events completely conserved in two or more plant species.

Conclusion: This study extends the range of plant taxa shown to have high levels of AS, confirms the importance of intron retention in plants, and demonstrates the utility of using ESTs from related species in order to identify novel and conserved AS events. The results also indicate that the frequency of AS in plants is comparable to that observed in mammals. Finally, our results highlight the importance of normalizing EST levels when estimating the frequency of alternative splicing.

Show MeSH