Characterization of Ghrelin O-Acyltransferase (GOAT) in goldfish (Carassius auratus)

Abstract

Ghrelin is the only known hormone posttranslationally modified with an acylation. This modification is crucial for most of ghrelin’s physiological effects and is catalyzed by the polytopic enzyme ghrelin O-acyltransferase (GOAT). The aim of this study was to characterize GOAT in a teleost model, goldfish (Carassius auratus). First, the full-length cDNA sequence was obtained by RT-PCR and rapid amplification of cDNA ends methods. Two highly homologous cDNAs of 1491 and 1413 bp, respectively, named goat-V1 and goat-V2 were identified. Deduced protein sequences (393 and 367 amino acids, respectively) are predicted to present 11 and 9 transmembrane regions, respectively, and both contain two conserved key residues proposed to be involved in catalysis: asparagine 273 and histidine 304. RT-qPCR revealed that both forms of goat mRNAs show a similar widespread tissue distribution, with the highest expression in the gastrointestinal tract and gonads and less but considerable expression in brain, pituitary, liver and adipose tissue. Immunostaining of intestinal sections showed the presence of GOAT immunoreactive cells in the intestinal mucosa, some of which colocalize with ghrelin. Using an in vitro approach, we observed that acylated ghrelin downregulates GOAT gene and protein levels in cultured intestine in a time-dependent manner. Finally, we found a rhythmic oscillation of goat mRNA expression in the hypothalamus, pituitary and intestinal bulb of goldfish fed at midday, but not at midnight. Together, these findings report novel data characterizing GOAT, and offer new information about the ghrelinergic system in fish.

Nesfatin-1 suppresses fish reproductive axis and gonadal steroidogenesis

“…iScript cDNA synthesis kit (Bio-Rad) was used for cDNA synthesis. Quantitative PCR (qPCR) was used as described earlier (Blanco et al 2017, Rajeswari et al 2019 for mRNA expression analysis employing the Universal SYBR Green Master Mix (Bio-Rad) and the CFX Connect Real-Time PCR Detection System (Bio-Rad). Primer validation and efficiency of primers were confirmed for each primer set as described earlier (Gonzalez et al 2012a).…”

Ghrelin enhances food intake and carbohydrate oxidation in a nitric oxide dependent manner

“…2015; Gutierrez et al, 2008; Romero et al, 2010; Mohan and Unniappan, 2013). GOAT is conserved across a variety of vertebrate, including mice ( Mus musculus ), rats ( Rattus norvegicus ), domestic chickens ( Gallus domesticus ) and goldfish ( Carassius auratus ) (Gutierrez et al, 2008; Seim et al, 2016; Blanco et al, 2017). Furthermore it appears that a truncated form of GHSR1a, GHSR1b, is also cleaved from the same gene and has been reported to reduce cell surface GHSR1a availability via heterodimerization (Chow et al, 2012).…”

Implicación de la MRAP2s en el sistema de melanocortinas de peces usando como modelos a la lubina europea y el pez cebra

“…Although the data seem to indicate that the (Kaiya et al 2005;Unniappan and Peter 2005), suggesting that ghrelin is present in the brain only in low quantities. By contrast, GOAT, the enzyme acylating ghrelin, is widely synthesized in the brain and its production is related to energy status (Blanco et al 2017;Hatef et al 2015). The role of GOAT in the postranscriptional processing of central ghrelin is uncertain and other roles including lipid acyl transferase cannot be ruled out.…”