Cloning and spatiotemporal expression of pepsinogen and gastric proton pump genes from mandarin fish (Siniperca chuatsi) during early ontogeny.

A fully developed stomach, characterized by the secretion of pepsinogens and chlorhydric acid, is vital for digestion and survival of fish larvae. To further understand the functional development of the stomach of mandarin fish (Siniperca chuatsi) during early ontogeny, the temporal and spatial expression of pepsinogens (PG A1, A2 and C), as well as proton pump genes were analyzed in the stomach from 0 to 40 days post-hatch (dph) by reverse transcription polymerase chain reaction (RT-PCR) and in situ hybridization (ISH) techniques. Pepsinogen C cDNA was firstly cloned with a full length of 1,557 bp, which contained a 37-bp 5'-untranslated region (UTR), an open reading frame of 1,164 bp encoding a polypeptide of 387 amino acids (aa) residues and a 356-bp 3'-UTR. RT-PCR analysis revealed a sequential expression mode of three pepsinogens (PG A1, A2 and C) along the ontogeny of the stomach in mandarin fish. Pepsinogen A1 was firstly detected at 4 dph (84 degree-days, dd) ahead of the appearance of gastric glands; pepsinogen A2 appeared at 12 dph (252 dd) and became the predominant form in the stomach after 19 dph (399 dd); pepsinogen C was the latest expressed gene at 14 dph (294 dd). Expression of proton pump at 12 dph (252 dd), coinciding with the expression time of pepsinogen A2 showed an excellent coordinated transcription mode between proton pump and pepsinogens. ISH analysis located the expression of three pepsinogens and α subunit of proton pump in the same gastric gland cells, which confirmed that they belonged to oxynticopeptic cells. In addition, oxynticopeptic cells developed and increased gradually from 14 to 40 dph. No transcripts of pepsinogens or proton pump were detected in surface mucous cells and mucous neck cells of the gastric mucosa. Our results implied the functional development of stomach in mandarin fish was closely related to pepsinogens expression.