Cloning and deorphanization of three inotocin (insect oxytocin/vasopressin-like) receptors and their ligand from the tick Ixodes scapularis
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Cloning and deorphanization of three inotocin (insect oxytocin/vasopressin-like) receptors and their ligand from the tick Ixodes scapularis. / Hauser, Frank; Al-Ribaty, Tara; Stebegg, Marisa; Thygesen, Gedske; Grimmelikhuijzen, Cornelis J.P.
In: Biochemical and Biophysical Research Communications, Vol. 680, 2023, p. 34-41.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - Cloning and deorphanization of three inotocin (insect oxytocin/vasopressin-like) receptors and their ligand from the tick Ixodes scapularis
AU - Hauser, Frank
AU - Al-Ribaty, Tara
AU - Stebegg, Marisa
AU - Thygesen, Gedske
AU - Grimmelikhuijzen, Cornelis J.P.
N1 - Publisher Copyright: © 2023 The Authors
PY - 2023
Y1 - 2023
N2 - Many insects produce the cyclic neuropeptide inotocin (CLITNCPRGamide), which is the insect orthologue of the mammalian neuropeptides oxytocin and vasopressin. These insects also have one inotocin G protein-coupled receptor (GPCR), which is the orthologue of the mammalian oxytocin and vasopressin receptors. The tick Ixodes scapularis belongs to the subphylum Chelicerata, an arthropod taxon different from insects, to which also spiders, scorpions, and mites belong. I. scapularis is an ectoparasite and a health risk for humans, because it transfers pathogenic microorganisms to its human host during a blood meal, thereby causing serious neurological diseases, among them Lyme disease and tick-borne encephalitis (TBE). By annotating the genomic sequence of I. scapularis, we previously found one presumed tick inotocin preprohormone gene and, in contrast to insects, three genes coding for presumed inotocin GPCRs. We now find that these GPCR genes cluster on one genomic contig, suggesting that they originated by recent gene duplications. Closely located on the same contig are also four adipokinetic hormone/corazonin-related peptide (ACP) GPCR genes, and one crustacean cardioactive peptide (CCAP) GPCR gene, suggesting evolutionary relationships. These evolutionary relationships are confirmed by phylogenetic tree analyses of their gene products. We also cloned the tick inotocin preprohormone, which has a structural organization closely resembling mammalian oxytocin and vasopressin preprohormones, including the presence of a conserved neurophysin sequence, having seven cystine bridges. This neurophysin sequence has two cystine-knot domains, but in contrast to mammalian neurophysins, the tick neurophysin contains a canonical prohormone convertase cleavage signal and a peptide C-terminal amidation sequence (GKR), suggesting cleavage into two biologically active cystine-knot peptides. This cleavage/amidation sequence occurs in neurophysins from most hard tick species, but not in other chelicerates. Mature tick inotocin is different from insect inotocin and has the sequence CFITNCPPGamide. Finally, we cloned and stably expressed the three tick inotocin receptors in Chinese Hamster Ovary cells and found that each of them was activated by nanomolar concentrations of tick inotocin (EC50 for ITR1 = 1.6 × 10−8 M; EC50 for ITR2 = 5.8 × 10−9 M; EC50 for ITR3 = 9.3 × 10−9 M), thereby establishing that they are genuine tick inotocin receptors.
AB - Many insects produce the cyclic neuropeptide inotocin (CLITNCPRGamide), which is the insect orthologue of the mammalian neuropeptides oxytocin and vasopressin. These insects also have one inotocin G protein-coupled receptor (GPCR), which is the orthologue of the mammalian oxytocin and vasopressin receptors. The tick Ixodes scapularis belongs to the subphylum Chelicerata, an arthropod taxon different from insects, to which also spiders, scorpions, and mites belong. I. scapularis is an ectoparasite and a health risk for humans, because it transfers pathogenic microorganisms to its human host during a blood meal, thereby causing serious neurological diseases, among them Lyme disease and tick-borne encephalitis (TBE). By annotating the genomic sequence of I. scapularis, we previously found one presumed tick inotocin preprohormone gene and, in contrast to insects, three genes coding for presumed inotocin GPCRs. We now find that these GPCR genes cluster on one genomic contig, suggesting that they originated by recent gene duplications. Closely located on the same contig are also four adipokinetic hormone/corazonin-related peptide (ACP) GPCR genes, and one crustacean cardioactive peptide (CCAP) GPCR gene, suggesting evolutionary relationships. These evolutionary relationships are confirmed by phylogenetic tree analyses of their gene products. We also cloned the tick inotocin preprohormone, which has a structural organization closely resembling mammalian oxytocin and vasopressin preprohormones, including the presence of a conserved neurophysin sequence, having seven cystine bridges. This neurophysin sequence has two cystine-knot domains, but in contrast to mammalian neurophysins, the tick neurophysin contains a canonical prohormone convertase cleavage signal and a peptide C-terminal amidation sequence (GKR), suggesting cleavage into two biologically active cystine-knot peptides. This cleavage/amidation sequence occurs in neurophysins from most hard tick species, but not in other chelicerates. Mature tick inotocin is different from insect inotocin and has the sequence CFITNCPPGamide. Finally, we cloned and stably expressed the three tick inotocin receptors in Chinese Hamster Ovary cells and found that each of them was activated by nanomolar concentrations of tick inotocin (EC50 for ITR1 = 1.6 × 10−8 M; EC50 for ITR2 = 5.8 × 10−9 M; EC50 for ITR3 = 9.3 × 10−9 M), thereby establishing that they are genuine tick inotocin receptors.
KW - Cystine-knot
KW - Evolution
KW - GPCR
KW - Neurophysin
KW - Oxytocin
KW - Vasopressin
U2 - 10.1016/j.bbrc.2023.09.009
DO - 10.1016/j.bbrc.2023.09.009
M3 - Journal article
C2 - 37716155
AN - SCOPUS:85171140676
VL - 680
SP - 34
EP - 41
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
SN - 0006-291X
ER -
ID: 367901816