Characterization of a third ghrelin receptor, GHS-R3a, in channel catfish reveals novel expression patterns and a high affinity for homologous ligand

Brian C Small; Sylvie M A Quiniou; Hiroyuki Kaiya; Jacob W Bledsoe; Bryan Musungu

doi:10.1016/j.cbpa.2018.11.013

Characterization of a third ghrelin receptor, GHS-R3a, in channel catfish reveals novel expression patterns and a high affinity for homologous ligand

Comp Biochem Physiol A Mol Integr Physiol. 2019 Mar:229:1-9. doi: 10.1016/j.cbpa.2018.11.013. Epub 2018 Nov 22.

Authors

Brian C Small¹, Sylvie M A Quiniou², Hiroyuki Kaiya³, Jacob W Bledsoe⁴, Bryan Musungu²

Affiliations

¹ Department of Fish and Wildlife Sciences, Aquaculture Research Institute, University of Idaho, 3059F National Fish Hatchery Road, Hagerman, ID 83332, United States. Electronic address: bcsmall@uidaho.edu.
² USDA/ARS Warmwater Aquaculture Research Unit, Thad Cochran National Warmwater Aquaculture Center, P.O. Box 38, Stoneville, MS 38776, United States.
³ Department of Biochemistry, National Cerebral and Cardiovascular Center Research Institute, Suita, Osaka 565-8565, Japan.
⁴ Department of Fish and Wildlife Sciences, Aquaculture Research Institute, University of Idaho, 3059F National Fish Hatchery Road, Hagerman, ID 83332, United States.

PMID: 30471349
DOI: 10.1016/j.cbpa.2018.11.013

Abstract

A novel third channel catfish growth hormone secretagogue (ghrelin) receptor, GHS-R3a, gene was characterized. Identification and analysis of the genomic organization of channel catfish GHS-R3a revealed differences in exon/intron structure relative to the previously published GHS-R1a and GHS-R2a sequences. Amino acid sequence alignment of catfish GHS-R3a with -R1a and -R2a revealed 48 and 52% sequence identity, respectively. Phylogenetic analysis predicted a new clade of GHS-R3a receptors found only in fish, with representation in the teleost infradivisions Osteoglossomorpha, Clupeomorpha, and Euteleostei. In functional analyses, homologous catfish ghrelin increased intracellular Ca²⁺ concentration in human embryonic kidney (HEK) 293 cells stably expressing catfish GHS-R3a. On the contrary, intracellular Ca²⁺ concentration was unaffected by treatment with the synthetic growth hormone secretagogues GHRP-6 and hexarelin. Realtime PCR results indicated high expression of GHS-R3a in the brain and gonads, demonstrating tissue specificity among the catfish GHS-Rs. The effects of fasting and refeeding on all three ghrelin receptors were evaluated in catfish brain, pituitary, stomach, and Brockmann bodies. Most notably, GHS-R3a was the only receptor observed to significantly increase (2.9-6.3-fold) in brain, pituitary, and stomach within 4 days of fasting (P < .05). Stomach GHS-R1a also increased (P < .05) after 4 days; however, GHS-R2a was only elevated in brain and pituitary after refeeding for 1 week. Expression of all three ghrelin receptors were elevated (P < .05) in the Brockmann bodies after 2 weeks of fasting and returned to prefasting levels following refeeding. Together with the previously published characterization of GHS-R1a and -R2a, these results establish three ghrelin receptors, each altered by energy state, in channel catfish and add to the growing body of information on GHS-R evolution and function.

Keywords: Catfish; Expression; Fasting; Function; Gene organization; Ghrelin; Growth hormone secretagogue; Sequence.

Publication types

Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Amino Acid Sequence
Animals
Base Sequence
DNA, Complementary / genetics
Dose-Response Relationship, Drug
Fasting
Fish Proteins / metabolism*
HEK293 Cells
Homeostasis
Humans
Ictaluridae / metabolism*
Ligands
Oligopeptides / administration & dosage
Oligopeptides / pharmacology
RNA, Messenger / genetics
Receptors, Ghrelin / antagonists & inhibitors
Receptors, Ghrelin / genetics
Receptors, Ghrelin / metabolism*
Sequence Homology, Amino Acid

Substances

DNA, Complementary
Fish Proteins
GHRP-6, Lys(3)-
Ligands
Oligopeptides
RNA, Messenger
Receptors, Ghrelin