Comparative genomic analysis of Drosophila melanogaster and vector mosquito developmental genes

Abstract

Genome sequencing projects have presented the opportunity for analysis of developmental genes in three vector mosquito species: Aedes aegypti, Culex quinquefasciatus, and Anopheles gambiae. A comparative genomic analysis of developmental genes in Drosophila melanogaster and these three important vectors of human disease was performed in this investigation. While the study was comprehensive, special emphasis centered on genes that 1) are components of developmental signaling pathways, 2) regulate fundamental developmental processes, 3) are critical for the development of tissues of vector importance, 4) function in developmental processes known to have diverged within insects, and 5) encode microRNAs (miRNAs) that regulate developmental transcripts in Drosophila. While most fruit fly developmental genes are conserved in the three vector mosquito species, several genes known to be critical for Drosophila development were not identified in one or more mosquito genomes. In other cases, mosquito lineage-specific gene gains with respect to D. melanogaster were noted. Sequence analyses also revealed that numerous repetitive sequences are a common structural feature of Drosophila and mosquito developmental genes. Finally, analysis of predicted miRNA binding sites in fruit fly and mosquito developmental genes suggests that the repertoire of developmental genes targeted by miRNAs is species-specific. The results of this study provide insight into the evolution of developmental genes and processes in dipterans and other arthropods, serve as a resource for those pursuing analysis of mosquito development, and will promote the design and refinement of functional analysis experiments.

Figure 1. Wnt Signaling Pathway Components in Mosquitoes.

The canonical Wnt pathway is summarized in this figure. Wnt binds its receptor Fz, which activates Dsh, an inhibitor of GSK-3. GSK-3 normally prevents dissociation of β-catenin from APC; when GSK-3 is inhibited, β-catenin enters the nucleus and regulates transcription (reviewed in [28]). Analysis of pathway members uncovered mosquito lineage specific changes in the number of orthologous sequences for genes encoding various Wnt pathway components in mosquitoes. With respect to D. melanogaster, mosquito lineage specific gene absences (−), as well as increases in the number of orthologues (+) were noted for particular pathway members. Details are provided in the text.

Figure 2. Evolutionary relationships of fz genes.

Relationships among orthologous mosquito and Drosophila Fz (Fz-1) proteins were inferred using the Neighbor-Joining method. The gene ID and the species name (5 letters) are shown for the orthologs. The optimal tree (the sum of branch length = 2.675) is shown. The percentage values of replicate trees in which the associated taxa clustered together following bootstrap testing (1000 replicates) are shown next to the branches. The tree is drawn to scale (shown below the tree), with branch lengths in the same units as those of the evolutionary distances used to infer the phylogeny. The distance scale is in units of the number of amino acid substitutions per site. The phylogenetic branching suggests that there could be an ancestral as well as a modern origin of mosquito fz genes in relation to Drosophila.

Figure 3. Phylogenetic relationships of FGF receptor genes.

D. melanogaster has two FGF receptors, Htl and Btl. A Neighbor-Joining tree of Htl and Btl proteins among mosquito and Drosophila species is shown. The gene ID and the species name (5 letters) are indicated for the orthologs. The optimal tree (the sum of branch length = 2.022) is shown. The percentage values of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) are shown next to the branches. The tree is drawn to scale (shown below the tree), with branch lengths in the same units as those of the evolutionary distances used to infer the phylogeny. The distance scale is in units of the number of amino acid substitutions per site. The results of these analyses indicate that mosquito FGF receptors are htl orthologs, and mosquitoes lack btl.

Figure 4. Regulation of cell death in dipterans.

In D. melanogaster, apoptosis is induced by the proapoptotic proteins Grim, Rpr, and Hid, which antagonize the function of dIap, thereby preventing it from inhibiting Dronc. When Dronc is active, DrICE, the main effector caspase of apoptosis in flies, is activated (reviewed in [67], [68]). Although many key components of the Drosophila apoptosis pathway are conserved in mosquitoes, mosquitoes lack orthologs of several key regulators of apoptosis (denoted by a – sign). In mosquitoes, Michelob_X is believed to function as the missing Iap antagonist . Additional details and discussion are provided in the text.

Figure 5. Cuticle Gene Phylogenetic Relationships.

Evolutionary relationships of adult cuticle protein genes (one-to-one orthologs) among mosquito and Drosophila species were inferred using the Neighbor-Joining method. The gene IDs and the species names (5 letters) are shown for the orthologs. The optimal tree (the sum of branch length = 0.842) is shown. The percentage values of replicate trees in which the associated taxa clustered together in bootstrap testing (1000 replicates) are shown next to the branches. The tree is drawn to scale (shown below the tree), with branch lengths in the same units as those of the evolutionary distances used to infer the phylogeny. The distance scale is in units of the number of amino acid substitutions per site. Although fruit fly and mosquito cuticle proteins are a highly conserved family of proteins, the one-to-one orthologs show lineage specific phylogenetic groupings among the 12 fruit fly and three mosquito species.

Figure 6. Targets of miRNAs in mosquito developmental genes.

The number of predicted developmental gene targets of miRNAs vary in number in D. melanogaster, A. aegypti, and A. gambiae. These results suggest that the repertoire of miRNA developmental gene targets may be species specific.