Multiple Vitellogenins (Vgs) in Mosquitofish (Gambusia affinis): Identification and Characterization of Three Functional Vg Genes and Their Circulating and Yolk Protein Products1

Abstract

The objectives of this study were to characterize multiple forms of vitellogenin (Vg) in mosquitofish (Gambusia affinis) and to discover the fate of each Vg during its processing into product yolk proteins. Two Vg preparations, with apparent masses of 600 kDa (600 Vg) and 400 kDa (400 Vg), were isolated from the plasma of fish treated with estradiol-17beta (E(2)) by various chromatographic procedures. Immunological analyses verified the presence of two different Vg proteins (600 VgA and 600 VgB) in the 600 Vg preparation and of a single protein in the 400 Vg preparation. Three major yolk proteins (Yps) with apparent masses of 560, 400, and 28 kDa were observed in extracts of ovarian follicles from vitellogenic females. Immunological analyses demonstrated that the 400 Vg underwent no change in native mass after being incorporated into oocytes. The 600 Vgs gave rise to a 28 kDa beta'-component and a native 560 kDa Yp, which was heterodimeric in structure, consisting of two types of complexes between phosvitin (Pv) and lipovitellin (Lv) heavy- and light-chains. Full-length cDNAs encoding the 600 VgA, 600 VgB, and 400 Vg were isolated from a liver cDNA library of E(2) treated fish. Similar to the zebrafish vg3 gene, the 400 Vg cDNA lacked a Pv domain and was classified as an incomplete or phosvitinless (C-type) Vg. The deduced primary structures of 600 VgA and 600 VgB were complete, and these were categorized as type A and type B Vgs, respectively, according to our recent classification scheme. This is the first report on the characterization of three functional Vg genes and their circulating and yolk protein products in any vertebrate species.

Vitellogenesis and choriogenesis in fishes

“…To date, the complete Vtg (VtgA and/or VtgB) and incomplete Vtg (VtgC) have been successfully isolated and purified as two different proteins with distinct molecular weights and antigenicity in various fish species, including tilapia (O. aureus [85] and O. mossambicus [86]), medaka [87], Japanese goby Acanthogobius flavimanus [88] and Sakhalin taimen [89]. In addition, three distinct types of Vtg have been purified or detected in mosquitofish Gambusia affinis [90], red seabream Pagrus major [91] and grey mullet [92]. Two or three types of Vtgs have been detected as proteins with distinct antigenicity in various existing fish species [66].…”

Biochemical and Immunochemical Characterization of Two Discrete Vitellogenin Proteins and Their Derived Lipovitellins in the Inshore Hagfish (Eptatretus burgeri)

“…As none of the above features fit to the aforementioned Vtg-related intermediate bands (bands migrating between ~62 to ~43 kDa), they might be proteolytic variants other than typical YP classes. A possible complex peptide composed of β′-c and Pv appeared to be present in the catshark (Yamane et al, 2013), while the presence of other YP variants with variable combinations of YP domains has been found in several fishes (Sawaguchi et al, 2005;Amano et al, 2007b). Thus, fishes may differ in the mode of Vtg cleavage, giving rise to a variety of Vtgderived YP peptides.…”

“…Prior to its deposition in yolk granules, Vtg undergoes limited proteolytic cleavage, giving rise to several classes of YPs. In teleosts, Vtg-derived YPs are typically designated as lipovitellin (Lv), phosvitin (Pv) and β′-component (β′-c) (Matsubara and Sawano, 1995;Hiramatsu and Hara, 1996;Hiramatsu et al, 2002b, c), while some other YP variants (e.g., Cterminal peptide and Lv-Pv complexes) are also evident Sawaguchi et al, 2005;Amano et al, 2007b).…”

“…Of the teleosts presenting three vtg transcripts, all corresponding Vtg proteins (VtgAa, VtgAb, and VtgC) have been identified in some species, including white perch, Morone americana (Hiramatsu et al, 2002c), mosquitofish, Gambusia affinis (Sawaguchi et al, 2005), red seabream, Pagrus major (Sawaguchi et al, 2006), and grey mullet, Mugil cephalus (Amano et al, 2007a).…”

Effects of adrenergic agents on the expression of zebrafish (Danio rerio) vitellogenin Ao1

“…With this assumption, heart tissue has often been overlooked to test for vtg mRNA levels in previous reports on medaka vtgAb, zebrafish vtgAo1, vtgC or many other newly cloned fish VTG transcripts (Wang et al, 2000;Islinger et al, 2003;Van der Ven et al, 2003;Tong et al, 2004;Wang et al, 2005;Jung et al, 2005;Sawaguchi et al, 2005;Mikawa et al, 2006). One exception is the report by Mikawa et al (2006) that indicated that vtgAe1 expression is not expressed in the heart tissue of the Japanese conger.…”