Fig 1.
Juvenile hormone III and cholesterol bio-synthesis pathways in insects, modified from Bellés et al.
[38].
Table 1.
Comparison of transcriptomes from different tick species, sexes, tissues and sequencing methods.
Fig 2.
Sequences alignment of farnesyl diphosphate synthase (FPPS2) from Bombyx mori and Drosophila melanogaster with contig 9824 and XP_002408650 from I. scapularis transcriptome and genome.
Farnesyl diphosphate synthase (FPPS2) described from Bombyx mori and Drosophila melanogaster aligned with contig 9824 from the I. scapularis transcriptome and FPPS (XP_002408650) from the I. scapularis genome. The boxed regions are the seven prenyltransferase conserved regions previously identified by Koyoma et al. [50]. “X” above the amino acids indicates position of Asp residues within the two aspartate-rich domains. Below the sequence alignment is the conservation panel which is measured as a numerical index (9–0) reflecting the conservation of physicochemical properties in the alignment. * (asterisk) denotes the highest identity score (identical residues in all species) followed by a score of 9 for the next most conserved group of residues containing substitutions by amino acids included in the same physicochemical class as described by Livingstone and Barton [52].
Table 2.
Longest contig from the synganglion transcriptomes of D. variabilis, I. scapularis, and O. turicata with the lowest e-values for matches in the mevalonate pathway (leading to the synthesis of juvenile hormone III) in insects1.
Fig 3.
Phylogenetic tree comparing the P450 CYP15A1 found in D. punctata and S. gregaria to P450s found in I. scapularis genome.
Phylogenetic tree comparing the P450s CYP15A1 known in D. punctata (AAS13464) and S. gregaria (HQ634703) to add the C10,11 epoxide to methyl farnesoate to make JH III, to the P450s in the I. scapularis genome with the top BLASTp matches (lowest e-values) to the D. punctata CYP15A1 and which had the longest length in base pairs. The 20 full length CYP messages with the lowest e-values and maximum length available from I. scapularis were not in the CYP15A1 family (in a different clade) as shown. The optimal neighbor-joining phylogenic tree was constructed by the Molecular Evolutionary Genetics Analysis (MEGA) program. All of the accession numbers labeled with XPs are CYP messages from the I. scapularis genome. The recombinant expressed CYP15A1s from D. punctata and S. gregaria are labeled insect JH epoxidase. Percent identity was determined by BLASTp. The highest identity message XP_002410454 shares 28% identity with D. punctata and 30% identity with S. gregaria. No members of the Cyp15A family were found in the I. scapularis genome [48]. An alignment of the closest sequence XP002410454 to the CYP15A1 family is shown in Fig 4.
Fig 4.
Sequence alignment of XP_002410454 from the I. scapularis genome with CYP15A1 from D. punctata AAS13464 and S. gregaria HQ634703.
Sequence alignment (pairwise) for XP_002410454, the top BLAST match from the I. scapularis genome to that of the published sequences (GenBank) for CYP15A1 from D. punctata AAS13464 [23] and S. gregaria HQ634703 [59]. The JH epoxidase is a member of the CYP15A1 family and has been cloned and characterized from the corpora allata of D. punctata and S. gregaria. It has been demonstrated this enzyme can add an epoxide to the C10,11 position of methyl farnesoate to produce JH III with high stereo selectivity (10R). All members of the family CYP15 share >40% identity at the amino acid level and >55% identity at the subfamily level [58]. The boxed sequence on the alignment is the signature heme-binding motif of the P450 (F**G***C*G). An * (asterisk) indicates positions which have a single, fully conserved residue. A colon indicates conservation between groups of strongly similar properties, scoring > 0.5 in the Gonnet PAM 250 matrix. A period indicates conservation between groups of weakly similar properties, scoring ≤ 0.5 in the Gonnet PAM 250 matrix. Based on our alignment, the I. scapularis sequence XP_002410454 only shares 28% identity with D. punctata and 30% identity with S. gregaria. Therefore XP_002410454 is not a member of the CYP15 gene family.
Table 3.
Longest contig from the synganglion transcriptomes of D. variabilis, I. scapularis, and O. turicata with the lowest e-values for matches in the JH branch in insects1.
Fig 5.
Alignment of methyltransferases from ticks with that of known JH methyltransferases from insects (based on published direct demonstration of function or by bioinformatics).
Contains the Gln-14 and Trp-120 residues which are important for farnesoic acid or JH acid interaction, and part of the SAM binding motif. Both Gln-14 and Trp-120 residues are labeled with + on the top of the sequence. The SAM binding motif and ligand interactions are marked with X on the top of the sequence. Selected methyltransferases with the maximum length in bps from the transcriptomes of D. variabilis, I. scapularis and O. trunicata were aligned with known insect JH methyl transferases from Bombyx mori (NP_001036901), Drosophila melanogaster (NP_609793), Tribolium castaneum (NP_001120783), and Aedes aegypti (XP_001651876).
Fig 6.
Farnesol dehydrogenase (AaSDR) from Aedes aegypti aligned with contig 5964 from the D. variabilis synganglion transcriptome.
Highlighted residues on both alignments show the conserved motifs that place them in the SDR family and the subfamily cP2. Below the sequence alignment is the conservation panel which is measured as a numerical index (9–0) reflecting the conservation of physicochemical properties in the alignment. * (asterisk) denotes the highest identity score (identical residues in all species) followed by a score of 9 for the next most conserved group of residues containing substitutions by amino acids included in the same physicochemical class as described by Livingstone and Barton [52].
Fig 7.
Farnesol dehydrogenase (AaSDR) from Aedes aegypti aligned with sequence EEC12752 from the I. scapularis genome.
Highlighted residues on both alignments show the conserved motifs that place them in the SDR family and the subfamily cP2. Below the sequence alignment is the conservation panel, which is measured as a numerical index (9–0) reflecting the conservation of physicochemical properties in the alignment. * (asterisk) denotes the highest identity score (identical residues in all species), followed by a score of 9 for the next most conserved group of residues containing substitutions by amino acids included in the same physicochemical class as described by Livingstone and Barton [52].
Fig 8.
Model for the endocrine regulation of vitellogenesis in D. variabilis and the potential role of the mevalonate-farnesal pathway.
Abbreviations in Fig: EDTH, hypothesized epidermal trophic hormone; Vg, vitellogenin; VgR, Vg receptor; 20-E, 20-hydroxyecdysone.