Characterization of a Novel α-Amidated Decapeptide Derived from Proopiomelanocortin-A in the Trout Pituitary1 (original) (raw)
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Endocrinology, 1997
Two complementary DNAs encoding distinct forms of POMC have been characterized in the trout pituitary. One of the POMC variants (POMC-A) possesses a C-terminal extension of 25 amino acids, which has no equivalent in other POMCs described to date. This C-terminal peptide contains three pairs of basic amino acids, suggesting that it may be the precursor of multiple processed peptides. In addition, the presence of a C-terminal glycine residue suggests that some of the processing products may be ␣-amidated. To characterize the molecular forms of the peptides generated from the C-terminal domain of trout POMC-A, we have developed specific antibodies against the C-terminal pentapeptide YHFQG and its ␣-amidated derivative YHFQ-NH 2 . Immunocytochemical labeling of pituitary sections with antibodies against YHFQ-NH 2 revealed the presence of numerous immunoreactive cells in the pars intermedia and the rostral pars distalis. In contrast, the antibodies against YHFQG produced only weak immunostaining. HPLC analysis combined with RIA detection revealed that extracts of the pars intermedia and pars distalis contain several peptides derived from the C-terminal extension of trout POMC-A, with the predominant molecular form exhibiting the same retention time as ALGERKYHFQ-NH 2 . Tryptic digestion of this major form produced a peptide that coeluted with YHFQ-NH 2 . These data indicate that the processing of the C-terminal extension of trout POMC-A generates several novel peptides including the decapeptide amide ALGERKYHFQ-NH 2 . (Endocrinology 138: 128 -137, 1997) FIG. 4. Microphotographs showing the distribution of YHFQG-immunoreactive cells in the pars intermedia (A) and the rostral pars distalis (B). Preincubation of the antiserum with synthetic YHFQG (10 Ϫ5 M) resulted in complete extinction of the immunoreaction in the pars intermedia (C) and rostral pars distalis (D). Scale bars ϭ 10 m.
Ontogeny of a novel decapeptide derived from POMC-A in the brain and pituitary of the rainbow trout
Developmental Brain Research, 2003
Trout POMC-A exhibits a unique C-terminal extension of 25-amino acids which is processed in the pituitary and hypothalamus to generate two novel decapeptides, EQWGREEGEE and ALGERKYHFQ-NH2. The fibers containing these two decapeptides are widely distributed in the brain, suggesting that they may exert neurotransmitter or neuromodulator activities. In the present study, we have investigated the ontogeny of the decapeptide EQWGREEGEE
Proopiomelanocortin (POMC)-related peptides in the brain of the rainbow trout, Salmo gairdneri
Peptides, 1989
investigated the presence of ACTH, et-MSH and 13-endorphin, three peptides which derive from the multifunctional precursor protein proopiomelanocortin (POMC) in the brain of the rainbow trout Salmo gairdneri. Using both the indirect immunofluorescence and peroxidase-antiperoxidase techniques, a discrete group of positive cells was identified in the hypothalamus, within the anterior part of the nucleus lateraiis tuberis, a-MSH-containing neurons represented the most abundant immunoreactive subpopulation. Coexistence of ct-MSH, ACTH and 13-endorphin was observed in the lateral part of the nucleus. ACTH-and 13-endorphin-containing cells were mainly distributed in the rostral and caudal regions of the nucleus. In the medial portion of the nucleus lateralis tuberis, numerous cells were only stained for a-MSH. Moderate to dense plexuses of immunoreactive fibers were observed in the ventral thalamus and the floor of the hypothalamus. Some of these fibers projected towards the pituitary. The concentrations of ACTH, et-MSH and 13-endorphin-like immunoreactivities were measured in microdissected brain regions by means of specific radioimmunoassays. Diencephalon, mesencephalon and medulla oblongata extracts gave dilution curves which were parallel to standard curves. The highest concentrations of POMC-derived peptides were found in the diencephalon (et-MSH: 4.28_+0.43 ng/mg prot.; ACTH: 1.08-+0.09 ng/mg prot.; 13-endorphin: 1.02-+ 0.1 ng/mg prot.), while lower concentrations were detected in the mesencephalon, medulla oblongatu and telencephalon. The present results demonstrate that various peptides derived from POMC coexist within the same cell bodies of the fish hypothalamus. Taken together, these data suggest that expression and processing of POMC in the fish brain is similar to that occurring in pituitary melanotrophs.
Fisheries Science, 2002
Several pituitary hormones, including adrenocorticotropin (ACTH), melanotropin (MSH) and b-endorphin, are generated from a common precursor protein, proopiomelanocortin (POMC). In fish, in addition to steroidogenesis of ACTH and melanogenesis of MSH, immunomodulating activity has been found in some POMC-related peptides. To investigate the functions of these peptides in the homologous system, it is necessary to establish a convenient detection method for the peptides. The present study aimed to establish a method for the detection of POMC-related peptides in bigeye tuna Thunnus obesus using a small amount of tissue sample, but not requiring peptide purification. We first determined the nucleotide sequence of tuna POMC cDNA. The cDNA was composed of 1084 base pairs (excluding the poly A tail) that encoded POMC consisting of 222 amino acids. We then fractionated an acid-acetone extract of one pituitary by reverse-phase high performance liquid chromatography (HPLC) and determined the molecular weight of each separated peptide by mass spectrometry. Consequently, we detected eight POMC-related peptides by comparing the values to the deduced amino acid sequence. Thus, the present study enabled the detection of POMC-related peptides from a small amount of tissue without the use of several purification steps.
General and Comparative Endocrinology, 1999
Proopiomelanocortin (POMC) cDNA was cloned from sea bass (Dicentrarchus labrax) pituitary gland. A 743 nucleotide sequence was obtained coding for the following sequences flanked by sets of proteolytic cleavage sites: ACTH (Ser 88-Met 127),-MSH (Ser 88-Gly 102), CLIP (Pro 106-Met 127),-LPH (Glu 131-Gln 208),-LPH (Glu 131-Ser 175),-MSH (Asp 159-Ser 175), and endorphin (Tyr 178-Gln 208). No region homologous to-MSH/joining peptide (a tetrapod POMC feature) was found. Amino acid sequence identity was high with other teleostean species considered (tilapia: 73%) and lower with elasmobranchs (dogfish: 42%). However, the presumed biologically active peptides were highly conserved within all species considered:-MSH (93-100%), ACTH (80-95%) and-endorphin (54-90%). Real-time PCR allowed us to quantify the expression of the POMC in different tissues of the sea bass: pituitary gland, liver, gonad and head kidney. No significant POMC expression was found in the integument. In pituitary gland, gonads, head kidney and liver, POMC expression was respectively, 1•26 10 10 , 2•67 10 5 , 2•06 10 4 and 1•67 10 4 copies/µg mRNA.
Journal of …, 2008
Nowadays, the studies of pituitary adenylate cyclase-activating polypeptide (PACAP)-related peptide (PRP) and PACAP in non-mammalian vertebrates, especially in fish, have paid attention mainly to the localization, cloning, and structural evolution of the peptides, but very little is known about its biological functions as growth-promoting factors in low vertebrates. In this work, we have cloned and characterized the PRP/PACAP cDNA from the commercially important North African catfish Clarias gariepinus. The sequence obtained agrees with the higher conservation of PACAP than of PRP peptide sequences. We have reported for the first time the recombinant expression of fish PRP and PACAP in mammalian cells and bacteria and also demonstrated that the growth rate of fish is enhanced by both PRP and PACAP recombinant peptides. The results obtained in vivo in three different fish species, catfish (C. gariepinus), tilapia (Oreochromis niloticus), and carp (Cyprinus carpio) support the finding that PACAP rather than PRP plays a primordial role in growth control in teleost fish. This finding could help to elucidate the neuroendocrine axis proposed to explain the hypothalamic regulation of growth in non-mammalian vertebrates.
General and Comparative Endocrinology, 2004
Several neurohormonal peptides of the gastrointestinal system of Wsh have been revealed by immunohistochemical methods. Among salmonids, the rainbow trout, Oncorhynchus mykiss (Walbaum) is the most studied species, whereas the informations about other species of the taxonomic group are lacking. The regional distribution and relative densities of cells belonging to the neuroendocrine system have been in this paper demonstrated in the gut of the brown trout, Salmo trutta Linnaeus. In the gastric mucosa, endocrine cells were detected, which were immunoreactive to bombesin-, gastrin-, and secretin-antisera. Endocrine cells containing gastrin-, bombesin-, cholecystokinin-8-, glucagon-, and leptin-like immunoreactivities were present in the pyloric caeca and intestine. The pancreatic endocrine islets contained glucagon-, and, possibly, secretin-like-immunoreactive endocrine cells, as well as a contingent of galanin-like-immunoreactive nerve Wbres. The exocrine pancreatic parenchyma showed bombesin-like-immunoreactive nerve Wbres. Within the tested regulatory peptides, bombesin and leptin were observed in both endocrine cells and nerve cell bodies and Wbres. Leptin was in addition detected in epithelial cells of the gastric glands. In the brown trout we have never observed any immunoreactivity to the VIP antiserum (either in the stomach or in the intestine). Some special structural patterns (in particular those ones related to galanin-and leptin-immunohistochemical data) have thus been detected for the Wrst time in the brown trout, and provide further data for a better knowledge of gut morpho-functional aspects in this economically important Wsh. 2004 Elsevier Inc. All rights reserved.