Insulin-like Growth Factor 1 (IGF-1): Orthologs and a Paralog in Rainbow Trout
This purpose of this project was to identify orthologs and paralogs of the rainbow trout IGF-1 gene.
Carl Ostberg
11-19-07
Figure 1: Nuclear sequence alignment
Figure 2: Neighbor-joining tree of nucleotide sequence orthologs
Figure 3: Protein sequence alignment
Figure 4: Neighbor-joining tree of protein sequence orthologs
3. IGF-1 Paralog in Rainbow Trout
Figure 5: Nuclear sequence alignment
Figure 6: Protein sequence alignment
4. Evolutionary relationship between IGF-1 and its paralog
Figure 7: Neighbor-joining tree of IGF-1 and its paralog in five different species
Disclaimer: This page was put together as a requirement for FISH 507A, University of Washington
IGF-1 is a polypeptide hormone that stimulates cellular division and growth. It is predominantly produced in the liver in response to growth hormone. Within humans, IGF-1 plays a significant role in growth and development during childhood and puberty. Within rainbow trout (Oncorhynchus mykiss), studies suggest that IGF-1 effects the following: reproduction and gonadal development (Loir 1999, Taylor 2007),
embryonic development (Li et al. 2007), osmoregulation (Liebert and Schreck 2006), nutritional regulation (Wilkinson et al. 2006, Li and Leatherland 2007), and cellular proliferation (Slagter et al. 2005). Although IGF-1 has multiple effects within rainbow trout, all these effects influence growth and development either directly or indirectly.
The IGF-1 gene is common among vertebrates, and has been isolated and sequenced across a diverse range of taxa (Rotwein et al. 1986, Shamblott and Chen 1992, Wu et al. 2006, Lu et al. 2007). The rainbow trout IGF-1 nucleotide sequence (Genebank accession number M95183) was used to generate nucleotide sequence alignments using several divergent taxa. Among fishes, nucleotide sequence alignments reveal IGF-1 orthologs are highly conserved (Figure 1). Inclusion of chicken, chimpanzee, and human nucleotide sequences with with fish sequences in Figure 1 resulted in a highly fragmented alignment with a completely different consensus sequence (data not shown). This result may be attributed to evolutionary divergence among introns and/or exons. Protein sequence alignments would be useful for identifying the nature of this difference (see below).
Figure 1. Nucleotide sequence alignments for IGF-1 orthologs in several species with the consensus sequence indicated. Several of the nucleotide sequences have been trimmed in order to condense the alignment. Genebank accession numbers are listed in parentheses.
Similarity among IGF-1 nucleotide sequence orthologs appears generally concordant with the phylogeny of fishes (Figure 2). However, it should be noted that Arctic charr, which should fall in with the other salmonids (rainbow trout, Atlantic salmon, coho salmon, and Chinook salmon) appeared to be more similar to Russian sturgeon. This could be due to poor sequence quality, a sequence isolated in Arctic charr other than IGF-1, or reflect evolutionary divergence of IGF-1 among salmonids. Further investigations of IGF-1 sequences in Arctic charr is required to resolve this issue.
Figure 2. Neighbor-joining tree of IGF-1 nucleotide sequence orthologs using the Jukes-Cantor genetic distance model with Russian sturgeon as the out group. Genebank accession numbers are listed in parentheses.
The rainbow trout IGF-1 protein sequence (Genebank accession number AAA49412) was used to generate protein sequence alignments using several divergent taxa. Protein sequence alignments indicate a high degree of homology among IGF-1 orthologs in fish, human, chicken, and chimpanzee (Figure 3). This suggests that the difference in nucleotide consensus sequence between fish only and the inclusion of human, chimpanzee, and chicken is most likely due to evolutionary divergence of introns. Further evidence for intron divergence is indicated by a fragmented nucleotide sequence, but a relatively complete protein alignment. Another observation is that there is extremely high homology among the protein sequences coding for the conserved domain (amino acids 66 - 134). Several other taxa divergent to vertebrates that are not known to express IGF-1 were also included (drosophila, nematode, neurospora, and trypanosoma). The protein alignments scores for these taxa were quite high (> 0.0001), indicating lack of homology between the vertebrate IGF-1 protein and the most closely related protein in drosophila, nematode, neurospora, and trypanosoma.
Figure 3. Protein sequence alignments for IGF-1 orthologs in several species with the consensus sequence indicated. The conserved domain for the protein sequence is an insulin-like growth factor occurring between amino acids 66 and 134 in the consensus sequence. Genebank accession numbers are listed in parentheses.
The evolutionary tree describing the relationship among IGF-1 proteins appears generally concordant with the phylogeny of the species included in the analysis (Figure 4). The evolutionary tree also indicates that the vertebrate IGF-1 protein sequences are divergent from the most closely related sequences in drosophila, nematode, neurospora, and trypanosoma. Within the vertebrates, there appears to be at least 3 lineages: Russian sturgeon, a primitive cartilaginous fish; terrestrial vertebrates; and aquatic vertebrates. A larger protein data set incorporating other cartilagenous fish plus species from several other taxa would provide interesting and useful results for partitioning out evolutionary relationships. Lastly, comparison of the nucleotide and protein sequence trees reveals similar phylogenies. These results suggests that IGF-1 may have evolved within vertebrates or the protein sequence and IGF-1 function may have diverged across several taxa.
Figure 4. Neighbor-joining tree of the IGF-1 protein orthologs using the Jukes-Cantor genetic distance model with Neurospora crassa as the out group. Genebank accession numbers are listed in parentheses.
3. IGF-1 Paralog in rainbow trout
IGF-2 is a paralog to IGF-1 (Rotwein 1991). Both IGF-1 and IGF-2 are expressed in a variety of tissues including liver, muscle, and plasma (Gabillard et al. 2003a). Similarly to IGF-1, the major function of IGF-2 is the promotion of growth and development. However, IGF-2 appears to be most active during embryonic development whereas IFG-1 is most active post-natal. In general, IGF-1 and IGF-2 perform similar functions, but are differentially expressed at different life-history stages (Gabillard et al. 2003a, Gabillard et al. 2003b). Within humans (Dull et al. 1984) and rainbow trout (Shamblott and Chen 1992) IGF-1 and IGF-2 are encoded by separate genes. A nucleotide sequence alignment between IGF-1 and IGF-2 in rainbow trout shows strong homology between 330 - 530 base pair region (Figure 5). The region of homology likely corresponds to a shared conserved domain.
Figure 5. The nucleotide sequence alignment for IGF-1 and its paralog (IGF-2) in rainbow trout with the consensus sequence indicated. Genebank accession numbers are listed.
The protein sequence alignment between IGF-1 and IGF-2 in rainbow trout indicates homology between the 50 - 117 amino acid region (Figure 6). This corresponds to 67 total amino acids and 201 total nucleotides, which was observed in Figure 5. The region of homology between the paralogs is the conserved domain which is an insulin-like growth factor. (Link to insulin-like growth factor conserved domain)
Figure 6. The protein sequence alignment for IGF-1 and its paralog (IGF-2) in rainbow trout with the consensus sequence indicated. The conserved domain for the protein sequence is an insulin-like growth factor occurring between amino acids 50 and 117 in the consensus sequence. Genebank accession numbers are listed.
4. Evolutionary relationship between IGF-1 and its paralog IGF-2
An evolutionary tree describing the relationship between IGF-1 and IGF-2 protein sequences suggests that these genes diverged early during vertebrate evolution (Figure 7). This is based on the observation that the chicken, human, rainbow trout, and zebra fish IGF-1 protein sequences were more closely related to each other than to the IGF-2 protein sequence within the same species. Interestingly, the IGF-1 and IGF-2 like proteins in drosophila were more similar to each other than they were to either protein in the other species.
Figure 7. Neighbor-joining tree of IGF-1 and its paralog, IGF-2, based on protein sequence data in five different species.
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