Isolation and partial characterization of a pair of prolactins released in vitro by the pituitary of a cichlid fish, Oreochromis mossambicus.

Abstract

The pituitary of the cichlid fish tilapia secretes two prolactins (PRLs) of molecular masses 20 kDa and 24 kDa. The 20-kDa PRL has an isoelectric point in the range of those of mammalian PRLs (pI 6.7), but the 24-kDa PRL is unusually basic (pI 8.7). Partial sequence information indicates that the PRLs are homologous but distinct proteins, differing by five amino acids within the first 29 NH2-terminal residues. Homology in the known region is higher with chum salmon PRL than with known mammalian PRLs. Reversed-phase HPLC permits isolation of these two PRLs and a single tilapia growth hormone from culture medium or from the pituitary in a single step. HPLC and radio-HPLC analysis of [3H]leucine pulse-chase experiments reveal that each PRL is secreted in vitro at remarkably high rates (21 pmol per gland per hr) and that the two PRLs are released in approximately equimolar amounts, suggesting the coordinate regulation of the secretion. Both PRLs exert characteristic PRL activity in that they prevent the loss of Na' from the plasma of hypophysectomized tilapia in fresh water.In teleost fishes, pituitary prolactin (PRL) production is confined to the rostral pars distalis (RPD). The RPD of the cichlid fish tilapia (Oreochromis mossambicus) consists almost entirely of PRL-secreting cells and accounts for almost one-third of the entire pituitary (1, 2). This regionalization of PRL cells differs from the scattered distribution of PRL cells in the pituitaries ofother vertebrates and has made the teleost pituitary a useful model for the study of PRL cell function.The aim of the investigations reported here was to make use of this organizational simplicity in order to isolate and characterize both stored and secreted PRL from the tilapia pituitary. This research direction was taken following reports that mammalian PRL might be modified during release (3-6) and because we thought that such modification might explain why tilapia PRL isolated previously exhibited anomalous electrophoretic behavior (7-9), even though its immunological and biological characteristics were similar to those of other vertebrate PRLs (10, 11).Our approach has been to culture the PRL-producing lobes and to isolate secreted proteins from the culture medium using reversed-phase HPLC.

Prolactin177, prolactin188 and prolactin receptor 2 in the pituitary of the euryhaline tilapia, Oreochromis mossambicus, are differentially osmosensitive

“…In tilapia, Prl 177 and Prl 188 are encoded by separate genes and differ in both amino acid number and composition, and consequently exhibit only 69% homology (Specker et al 1985, Yamaguchi et al 1988, Rentier-Delrue et al 1989. While the discovery of two distinct Prl molecules in tilapia suggested the evolution of distinct actions, no clear differences in osmoregulatory action have been demonstrated (Specker et al 1985). This issue has been recently revisited by Fiol et al (2009) Early studies have shown greater RPD and Prl cell size (Dharmamba & Nishioka 1968), greater Prl cell activity (Nagahama et al 1975), and greater hypophyseal Prl content (Nicoll et al 1981) in tilapia maintained in FW compared to tilapia maintained in SW. Later, Yoshikawa-Ebesu et al (1995) reported that hyposmotically induced Prl 177 and Prl 188 synthesis is more robust in RPDs of fish acclimated to FW than in RPDs of fish acclimated to SW.…”

“…In addition, recent evidence suggests that this difference in responsiveness may result from the diminished capability of (Specker et al 1985, Yamaguchi et al 1988, Rentier-Delrue et al 1989. While the existence of two distinct Prl molecules, designated Prl 177 and Prl 188 , suggests an evolution of differing actions (Chen et al 1994), a bioassay failed to identify any clear differences in their osmoregulatory activities (Specker et al 1985). On the other hand, it has been reported that Prl 177 exhibits somatotropic activity while Prl 188 does not (Shepherd et al 1997).…”

“…The tilapia pituitary releases two Prl molecules that differ in length and amino acid composition, and are encoded by separate genes (Specker et al 1985, Yamaguchi et al 1988, Rentier-Delrue et al 1989. While the existence of two distinct Prl molecules, designated Prl 177 and Prl 188 , suggests an evolution of differing actions (Chen et al 1994), a bioassay failed to identify any clear differences in their osmoregulatory activities (Specker et al 1985).…”

“…One likely explanation for this pattern is the fact that Prl cells from FW tilapia contain more stored Prl than those from SW fish (Nagahama et al 1975, Specker et al 1993 and, hence, exhibit a greater capacity to respond rapidly with hormone secretion when faced with a hyposmotic stimulus. In addition, recent evidence suggests that this difference in responsiveness may result from the diminished capability of (Specker et al 1985, Yamaguchi et al 1988, Rentier-Delrue et al 1989. While the existence of two distinct Prl molecules, designated Prl 177 and Prl 188 , suggests an evolution of differing actions (Chen et al 1994), a bioassay failed to identify any clear differences in their osmoregulatory activities (Specker et al 1985).…”

“…In tilapia, Prl 177 and Prl 188 are encoded by separate genes and differ in both amino acid number and composition, and consequently exhibit only 69% homology (Specker et al 1985, Yamaguchi et al 1988, Rentier-Delrue et al 1989. While the discovery of two distinct Prl molecules in tilapia suggested the evolution of distinct actions, no clear differences in osmoregulatory action have been demonstrated (Specker et al 1985). This issue has been recently revisited by Fiol et al (2009), who reported that Prl 177 and Prl 188 activate distinct downstream signalling pathways in transfected HEK293 cells expressing tilapia PrlRs.…”