Molecular cloning and characterisation of an isotocin paralogue ([V8] isotocin) in catfishes (superorder Ostariophysi): Origin traced likely to the fish‐specific whole genome duplication

Abstract

The present study reports the molecular cloning of a previously uncharacterised neurohypophyseal nonapeptide precursor cDNA in two catfish species: Heteropneustes fossilis and Clarias batrachus. The deduced nonapeptide is CYISNCPVG ([V8] isotocin), which has not been reported in any vertebrate till date. Phylogenetic and conserved synteny analyses showed the gene to have originated from the isotocin precursor (pro‐it) gene by fish‐specific whole genome duplication (3R). The two isotocin lineages have been designated as pro‐ita (new gene) and pro‐itb (conventional it gene). All teleost groups may not possess both pro‐ita and pro‐itb and the pattern of losses/retention was found to be lineage‐specific. Quantitative reverse transcriptase‐polymerase chain reaction studies showed the expression of the pro‐ita gene in the brain and ovary of H. fossilis. In situ hybridisation studies localised the pro‐ita transcripts in the nucleus preopticus of the hypothalamus and the follicular layer (theca‐granulosa) of oocytes, comprising tissues in which pro‐itb and vasotocin precursor (pro‐vt) mRNA expression was previously reported. The transcript levels varied with the reproductive stage and a high abundance was found in both brain and ovary during the breeding phase. The substitution of valine in place of isoleucine at the eighth position in Ita may have modified the ligand‐receptor interaction, leading to sub‐functionalisation and the retention of the gene in catfishes.

Reproductive roles of the vasopressin/oxytocin neuropeptide family in teleost fishes

“…In basal cephalochordates, such as the Florida lancelet, Branchiostoma floridae, and urochordates such as the sea vase, Ciona intestinalis, single nonapeptides, [Ile 4 ]-VP and Ciona (ciVP) have been reported (2)(3)(4). The presence of a single AVP family peptide extends to basal vertebrate agnathans, including lampreys such as the Japanese lamprey, Lethenteron japonicum, and hagfishes such as Eptatretus burgeri (5,6). It is believed that two rounds of whole genome duplication (2R WGD), one before and one after the separation of agnathans and gnathostomes (7), led to a duplication of the ancestral arginine vasotocin (avp) gene.…”

“…In teleost fishes, vasotocin (Avp) was originally isolated and characterized in pout, Gadus luscus (10), pollock, Polacchius virens (11), and European hake, Mercluccius mercluccius (12), and has since been identified in genomes of all teleost fishes studied to date (3,6). Avp occurs in all verebrates except for mammals, where it appears to have mutated to give rise to vasopressin (AVP) in which 3 Ile was substituted for 3 Phe (3,5,6). All AVP family members are basic nonapeptides due to a basic amino acid (AA) at position 8.…”

“…In contrast to the AVP family peptides, the duplicated nonapeptide gene appears to have accumulated more mutations giving rise to the OXT family (3,5,6). The OXT nonapeptides are characterized by a neutral AA at position 8 (Leu, Ile, Gln or Val).…”

“…While position 8 mutations are very common in cartilaginous fishes (19) specific Oxt family peptides have also been reported in Sarcopterygii, such as the Australian lungfish, Neoceratodus forsteri, which expresses [Phe 2 -Ile 3 ] OXT (20). Based on more recent genomic data, it has become evident that non-teleost Actinopterygians also express specific Oxt peptides (6). Indeed, in the spotted gar, Lepisosteus oculatus, which possess a pre-3R genome, two gene paralogues coding for Oxt were identified -one coding for a pro-[Ser 4 -Ile 8 ]-OXT with a long C-terminal (NCBI Accession No.…”

“…More recently however, the paradigm that teleost fishes, while encoding multiple gene loci for neutral nonapeptides in some cases, nevertheless exclusively express Oxt ([Ser 4 -Ile 8 ]-OXT) was challenged by the cloning and discovery of two different neutral Oxt family peptides in the Asian stinging catfish, Heteropneustes fossilis, and the walking catfish, Clarias batrachus (6). In both species, a conventional oxt gene coding for a [Ser 4 -Ile 8 ]-OXT and a second oxt gene coding for the novel [Ser 4 -Val 8 ]-OXT, coined sevatocin, are expressed in addition to a single avt gene (6). Like pro-[Ser 4 -Ile 8 ]-OXT, the peptide precursor encoding for [Ser 4 -Val 8 ]-OXT has a similar organization (Table 2), with an extended C-terminal and a Leu-rich region.…”