Limits...
RNA editing and alternative splicing of the insect nAChR subunit alpha6 transcript: evolutionary conservation, divergence and regulation.

Jin Y, Tian N, Cao J, Liang J, Yang Z, Lv J - BMC Evol. Biol. (2007)

Bottom Line: The occurrence of alternative splicing was found to be regulated in distinct modes and, in some cases, even correlated with RNA editing.On the basis of comparative analysis of orthologous nAChR alpha6 genes from different insects spanning ~300 million years of evolution, we have documented the existence, evolutionary conservation and divergence, and also regulation of RNA editing and alternative splicing.Phylogenetic analysis of RNA editing and alternative splicing, which can create a multitude of functionally distinct protein isoforms, might have a crucial role in the evolution of complex organisms beyond nucleotide and protein sequences.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Biochemistry, College of Life Sciences, Zhejiang University (Zijingang Campus), Hangzhou, Zhejiang, PR of China. jinyf@zju.edu.cn

ABSTRACT

Background: RNA editing and alternative splicing play an important role in expanding protein diversity and this is well illustrated in studies of nicotinic acetylcholine receptors (nAChRs).

Results: Here, we compare the RNA editing and alternative splicing of the nAChR alpha6 subunit genes from different insects spanning ~300 million years of evolution- Drosophila melanogaster, Anopheles gambiae, Bombyx mori, Tribolium castaneum and Apis mellifera. The conserved and species-specific A-to-I RNA editing occurred across all species except A. gambiae, which displayed extraordinarily short flanking intronic sequences. Interestingly, some A-to-I editing sites were a genomically encoded G in other species. A combination of the experimental data and computational analysis of orthologous alpha6 genes from different species indicated that RNA editing and alternative splicing predated at least the radiation of insect orders spanning ~300 million years of evolution; however, they might have been lost in some species during subsequent evolution. The occurrence of alternative splicing was found to be regulated in distinct modes and, in some cases, even correlated with RNA editing.

Conclusion: On the basis of comparative analysis of orthologous nAChR alpha6 genes from different insects spanning ~300 million years of evolution, we have documented the existence, evolutionary conservation and divergence, and also regulation of RNA editing and alternative splicing. Phylogenetic analysis of RNA editing and alternative splicing, which can create a multitude of functionally distinct protein isoforms, might have a crucial role in the evolution of complex organisms beyond nucleotide and protein sequences.

Show MeSH
The conserved and species-specific A-to-I RNA editing. Alignment of the homologous exon 5 genomic nucleotide (A) and amino acid (B) sequences of nAChR subunit alpha6 genes from D. melanogaster (Dme), A. gambiae (Aga), H. virescens (Hvi), B. mori (Bmo), T. castaneum (Tca) and A. mellifera (Ame). RNA editing of the nAChR subunit alpha6 genes from D. melanogaster, H. virescens and A. gambiae have been previously described [16, 17]. The editing sites from positions 1–14 (A) and amino acids (B) are shaded in red. Those sites constitutively G (A) and amino acids (B) are shaded in green at the editing sites. RNA editing sites, which were not evidently detected as a mixed sequence signal G and A, but revealed by sequencing of cDNA clones, are underlined.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC1919356&req=5

Figure 5: The conserved and species-specific A-to-I RNA editing. Alignment of the homologous exon 5 genomic nucleotide (A) and amino acid (B) sequences of nAChR subunit alpha6 genes from D. melanogaster (Dme), A. gambiae (Aga), H. virescens (Hvi), B. mori (Bmo), T. castaneum (Tca) and A. mellifera (Ame). RNA editing of the nAChR subunit alpha6 genes from D. melanogaster, H. virescens and A. gambiae have been previously described [16, 17]. The editing sites from positions 1–14 (A) and amino acids (B) are shaded in red. Those sites constitutively G (A) and amino acids (B) are shaded in green at the editing sites. RNA editing sites, which were not evidently detected as a mixed sequence signal G and A, but revealed by sequencing of cDNA clones, are underlined.

Mentions: Similar to substitution alternative splicing, RNA editing causes amino acid changes by substituting individual nucleotides. Although RNA A-to-I editing occurred in Drosophila transcripts, it was not present in the equivalent nAChR subunit of Anopheles [16,17]. To determine whether orthologous nAChR subunits were also RNA-edited in other insect species, we have subsequently analyzed the nAChR alpha6 genes from B. mori, T. castaneum and A. mellifera. Several sites were either determined as pure G signals, or as mixed sequence signals of G and A, while the nucleotide in the reference genomic DNA was A at these positions. We also checked for genetic variation in these alpha6 regions, using PCR amplification on silkworm genomic DNA, applying primers surrounding the equivalent regions and direct sequencing. As a result, it was revealed that the nucleotide in genomic DNA was adenosine at these positions, confirming that post-transcriptional modifications occur in this case. We have identified a total of ten A-to-I RNA editing sites within silkworm alpha6 transcripts, seven of which are located in exon 5 (Figure 5A). Sequence analysis of transcripts showed that six amino acids could be changed by seven possible A-to-G transitions in exon 5 (Figure 5B).


RNA editing and alternative splicing of the insect nAChR subunit alpha6 transcript: evolutionary conservation, divergence and regulation.

Jin Y, Tian N, Cao J, Liang J, Yang Z, Lv J - BMC Evol. Biol. (2007)

The conserved and species-specific A-to-I RNA editing. Alignment of the homologous exon 5 genomic nucleotide (A) and amino acid (B) sequences of nAChR subunit alpha6 genes from D. melanogaster (Dme), A. gambiae (Aga), H. virescens (Hvi), B. mori (Bmo), T. castaneum (Tca) and A. mellifera (Ame). RNA editing of the nAChR subunit alpha6 genes from D. melanogaster, H. virescens and A. gambiae have been previously described [16, 17]. The editing sites from positions 1–14 (A) and amino acids (B) are shaded in red. Those sites constitutively G (A) and amino acids (B) are shaded in green at the editing sites. RNA editing sites, which were not evidently detected as a mixed sequence signal G and A, but revealed by sequencing of cDNA clones, are underlined.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1919356&req=5

Figure 5: The conserved and species-specific A-to-I RNA editing. Alignment of the homologous exon 5 genomic nucleotide (A) and amino acid (B) sequences of nAChR subunit alpha6 genes from D. melanogaster (Dme), A. gambiae (Aga), H. virescens (Hvi), B. mori (Bmo), T. castaneum (Tca) and A. mellifera (Ame). RNA editing of the nAChR subunit alpha6 genes from D. melanogaster, H. virescens and A. gambiae have been previously described [16, 17]. The editing sites from positions 1–14 (A) and amino acids (B) are shaded in red. Those sites constitutively G (A) and amino acids (B) are shaded in green at the editing sites. RNA editing sites, which were not evidently detected as a mixed sequence signal G and A, but revealed by sequencing of cDNA clones, are underlined.
Mentions: Similar to substitution alternative splicing, RNA editing causes amino acid changes by substituting individual nucleotides. Although RNA A-to-I editing occurred in Drosophila transcripts, it was not present in the equivalent nAChR subunit of Anopheles [16,17]. To determine whether orthologous nAChR subunits were also RNA-edited in other insect species, we have subsequently analyzed the nAChR alpha6 genes from B. mori, T. castaneum and A. mellifera. Several sites were either determined as pure G signals, or as mixed sequence signals of G and A, while the nucleotide in the reference genomic DNA was A at these positions. We also checked for genetic variation in these alpha6 regions, using PCR amplification on silkworm genomic DNA, applying primers surrounding the equivalent regions and direct sequencing. As a result, it was revealed that the nucleotide in genomic DNA was adenosine at these positions, confirming that post-transcriptional modifications occur in this case. We have identified a total of ten A-to-I RNA editing sites within silkworm alpha6 transcripts, seven of which are located in exon 5 (Figure 5A). Sequence analysis of transcripts showed that six amino acids could be changed by seven possible A-to-G transitions in exon 5 (Figure 5B).

Bottom Line: The occurrence of alternative splicing was found to be regulated in distinct modes and, in some cases, even correlated with RNA editing.On the basis of comparative analysis of orthologous nAChR alpha6 genes from different insects spanning ~300 million years of evolution, we have documented the existence, evolutionary conservation and divergence, and also regulation of RNA editing and alternative splicing.Phylogenetic analysis of RNA editing and alternative splicing, which can create a multitude of functionally distinct protein isoforms, might have a crucial role in the evolution of complex organisms beyond nucleotide and protein sequences.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Biochemistry, College of Life Sciences, Zhejiang University (Zijingang Campus), Hangzhou, Zhejiang, PR of China. jinyf@zju.edu.cn

ABSTRACT

Background: RNA editing and alternative splicing play an important role in expanding protein diversity and this is well illustrated in studies of nicotinic acetylcholine receptors (nAChRs).

Results: Here, we compare the RNA editing and alternative splicing of the nAChR alpha6 subunit genes from different insects spanning ~300 million years of evolution- Drosophila melanogaster, Anopheles gambiae, Bombyx mori, Tribolium castaneum and Apis mellifera. The conserved and species-specific A-to-I RNA editing occurred across all species except A. gambiae, which displayed extraordinarily short flanking intronic sequences. Interestingly, some A-to-I editing sites were a genomically encoded G in other species. A combination of the experimental data and computational analysis of orthologous alpha6 genes from different species indicated that RNA editing and alternative splicing predated at least the radiation of insect orders spanning ~300 million years of evolution; however, they might have been lost in some species during subsequent evolution. The occurrence of alternative splicing was found to be regulated in distinct modes and, in some cases, even correlated with RNA editing.

Conclusion: On the basis of comparative analysis of orthologous nAChR alpha6 genes from different insects spanning ~300 million years of evolution, we have documented the existence, evolutionary conservation and divergence, and also regulation of RNA editing and alternative splicing. Phylogenetic analysis of RNA editing and alternative splicing, which can create a multitude of functionally distinct protein isoforms, might have a crucial role in the evolution of complex organisms beyond nucleotide and protein sequences.

Show MeSH