Atlantic SalmonHNF-3/forkhead: cDNA Sequence, Evolution, Expression, and Functional Analysis

Abstract

We report the isolation and characterization of a cDNA encoding an HNF-3 family member (as HNF-3) from Atlantic salmon (Salmo salar L). The important functional domains of HNF-3 proteins that have been characterized previously are revealed by segments of high identity along the alignment of the asHNF-3 with winged helix/forkhead amino acid sequences isolated from other species. A comparison of asHNF-3 cDNA and genomic DNA indicated that there were no introns present in the asHNF-3 gene. Expression of asHNF-3 protein in adult salmon tissues was not exclusive to liver but was also present in the pancreas and intestine. An RT-PCR analysis performed on salmon development showed that asHNF3 expression is detectable before gastrulation at the mid blastula transition stage. Functional analysis of the asHNF-3 protein using a characterized HNF-3 consensus binding site demonstrated that the protein can recognize and bind to specific HNF-3 consensus sequences. We also report the identification of a novel HNF3 binding site in the promoter of the Atlantic salmon transferrin gene.

Hepatocyte Nuclear Factors-1α, -1β, and -3β Expressed in the Gonad of Tilapia (Oreochromis mossambicus)

“…A similar result was reported for HNF-3 in Atlantic salmon (18). On the other hand, when only the winged helix/forkhead domain is consid- ered (Fig.…”

“…This suggests that differences in the forkhead family occurred before the divergence of tetrapods and teleosts. The strong conservation of the forkhead domain between piscine and mammalian species also illustrates the importance of the sequence defining the structure of the forkhead domain (18).…”

“…For example, while HNF-3␤ is expressed in chick liver, kidney, lung, muscle, and oviduct at a size of between 1.8 and 2.3 kb (32), it is not expressed in the kidney or muscle of mammals (3,33). Similarly, in Atlantic salmon, HNF-3␥ is expressed in the liver, intestine, and pancreas at a size of 3.6 kb, but is not expressed in the kidney, heart, ovary, spleen, or muscle (18). However, the results of semiquantitative RT-PCR show that in tilapia the IGF or HNF levels in the testis were not significantly different from the IGF and HNF levels in the ovary (Figs.…”

“…During mouse embryonic development, HNF-3␣ and HNF-3␤ are sequentially expressed in the defined endoderm, notochord, and floor plate of the neural tube, with HNF-3␤ being expressed first (5,16). The limited literature that deals with HNFs in bony fish focuses on the expression of HNFs during embryonic development or on their functional assays (17)(18)(19) similar to those used in mammals. However, the roles of HNFs in the gonadal physiology of mammals and fish are still not clear.…”

“…Furthermore, promoter regions of the IGF-I gene in tilapia (O. mossambicus) and chum salmon (Oncorhynchus keta) contain an HNF-1␣ binding site (22,23), and in combination with STAT5 after GH induction, HNF-1␣ can regulate the expression of the IGF-I gene (24). In Atlantic salmon, an HNF-3␤ binding site has been found in the IGF-II gene promoter regions, and this can activate the expression of IGF-II (Salmo salar) (18). Further, as in humans, where HNF-3␤ can transactivate IGF-I either directly or indirectly by activation of HNF-1␣ (25), recent evidence indicates that HNF-1␣ and HNF-3␤ can activate the promoter of salmon IGF-I (26) and IGF-II genes (17), respectively.…”