Nagasawa Kazue, Osugi Tomohiro, Suzuki Iwao, Itoh Naoki, Takahashi Keisuke G, Satake Honoo, Osada Makoto
Laboratory of Aquacultural Biology, Graduate School of Agricultural Science, Tohoku University, 1-1 Amamiya-Machi, Tsutsumidori, Aoba-Ku, Sendai, Miyagi 981 8555, Japan.
Suntory Institute for Bioorganic Research, 1-1-1 Wakayamadai, Shimamoto, Mishima, Osaka 618-8503, Japan.
Peptides. 2015 Sep;71:202-10. doi: 10.1016/j.peptides.2015.07.022. Epub 2015 Jul 31.
There is yet no firm experimental evidence that the evolutionary ancient gonadotropin-releasing hormone GnRH (i.e., GnRH1) also acts in invertebrate gametogenesis. The objective of this paper is to characterize candidate invGnRH peptides of Yesso scallop Patinopecten yessoensis (i.e., peptide identification, immunohistochemical localization, and immunoquantification) in order to reveal their bioactive form in bivalves. Using mass spectrometry (MS), we identified two invGnRH (py-GnRH) peptides from the scallop nerve ganglia: a precursor form of py-GnRH peptide (a non-amidated dodecapeptide; py-GnRH12aa-OH) and a mature py-GnRH peptide (an amidated undecapeptide; py-GnRH11aa-NH2). Immunohistochemical staining allowed the localization of both py-GnRH peptides in the neuronal cell bodies and fibers of the cerebral and pedal ganglia (CPG) and the visceral ganglion (VG). We found that the peptides showed a dimorphic distribution pattern. Notably, the broad distribution of mature py-GnRH in neuronal fibers elongating to peripheral organs suggests that it is multi-functional. Time-resolved fluorescent immunoassays (TR-FIA) enabled the quantification of each py-GnRH form in the single CPG or VG tissue obtained from one individual. In addition, we observed greater abundance of mature py-GnRH in VG compared with its level in CPG, suggesting that VG is the main producing organ of mature py-GnRH peptide and that py-GnRH may play a central regulatory role in neurons of scallops. Our study provides evidence, for the first time, for the presence of precursor and mature forms of invGnRH peptides in the nerve ganglia of an invertebrate.
目前尚无确凿的实验证据表明进化上古老的促性腺激素释放激素GnRH（即GnRH1）也在无脊椎动物配子发生中起作用。本文的目的是鉴定虾夷扇贝（Patinopecten yessoensis）的候选无脊椎动物GnRH肽（即肽鉴定、免疫组织化学定位和免疫定量），以揭示它们在双壳类动物中的生物活性形式。我们使用质谱（MS）从扇贝神经节中鉴定出两种无脊椎动物GnRH（py-GnRH）肽：一种py-GnRH肽的前体形式（一种非酰胺化的十二肽；py-GnRH12aa-OH）和一种成熟的py-GnRH肽（一种酰胺化的十一肽；py-GnRH11aa-NH2）。免疫组织化学染色使两种py-GnRH肽定位在脑足神经节（CPG）和内脏神经节（VG）的神经元细胞体和纤维中。我们发现这些肽呈现出双态分布模式。值得注意的是，成熟的py-GnRH在延伸至外周器官的神经纤维中的广泛分布表明它具有多种功能。时间分辨荧光免疫分析（TR-FIA）能够对从单个个体获得的单个CPG或VG组织中的每种py-GnRH形式进行定量。此外，我们观察到VG中成熟py-GnRH的丰度高于其在CPG中的水平，这表明VG是成熟py-GnRH肽的主要产生器官，并且py-GnRH可能在扇贝的神经元中发挥核心调节作用。我们的研究首次为无脊椎动物神经节中存在无脊椎动物GnRH肽的前体和成熟形式提供了证据。
