Dickinson Patsy S, Stemmler Elizabeth A, Barton Elizabeth E, Cashman Christopher R, Gardner Noah P, Rus Szymon, Brennan Henry R, McClintock Timothy S, Christie Andrew E
Department of Biology, Bowdoin College, 6500 College Station, Brunswick, ME 04011, USA.
Peptides. 2009 Feb;30(2):297-317. doi: 10.1016/j.peptides.2008.10.009. Epub 2008 Oct 21.
Recently, cDNAs encoding prepro-orcokinins were cloned from the crayfish Procambarus clarkii; these cDNAs encode multiple copies of four orcokinin isoforms as well as several other peptides. Using the translated open reading frames of the P. clarkii transcripts as queries, five ESTs encoding American lobster Homarus americanus orthologs were identified via BLAST analysis. From these clones, three cDNAs, each encoding one of two distinct prepro-hormones, were characterized. Predicted processing of the deduced prepro-hormones would generate 13 peptides, 12 of which are conserved between the 2 precursors: the orcokinins NFDEIDRSGFGFN (3 copies), NFDEIDRSGFGFH (2 copies) and NFDEIDRSGFGFV (2 copies), FDAFTTGFGHN (an orcomyotropin-related peptide), SSEDMDRLGFGFN, GDY((SO3))DVYPE, VYGPRDIANLY and SAE. Additionally, one of two longer peptides (GPIKVRFLSAIFIPIAAPARSSPQQDAAAGYTDGAPV or APARSSPQQDAAAGYTDGAPV) is predicted from each prepro-hormone. MALDI-FTMS analyses confirmed the presence of all predicted orcokinins, the orcomyotropin-related peptide, and three precursor-related peptides, SSEDMDRLGFGFN, GDYDVYPE (unsulfated) and VYGPRDIANLY, in H. americanus neural tissues. SAE and the longer, unshared peptides were not detected. Similar complements of peptides are predicted from P. clarkii transcripts; the majority of these were detected in its neural tissues with mass spectrometry. Truncated orcokinins not predicted from any precursor were also detected in both species. Consistent with previous studies in the crayfish Orconectes limosus, NFDEIDRSGFGFN increased mid-/hindgut motility in P. clarkii. Surprisingly, the same peptide, although native to H. americanus, did not affect gut motility in this species. Together, our results provide the framework for future investigations of the regulation and physiological function of orcokinins/orcokinin precursor-related peptides in astacideans.
最近，从小龙虾克氏原螯虾中克隆出了编码前激素原章鱼肽的cDNA；这些cDNA编码四种章鱼肽亚型的多个拷贝以及其他几种肽。以克氏原螯虾转录本的翻译开放阅读框为查询序列，通过BLAST分析鉴定出五个编码美洲龙虾同源物的EST。从这些克隆中，鉴定出三个cDNA，每个cDNA编码两种不同前激素原中的一种。预测推导的前激素原的加工过程会产生13种肽，其中12种在这两种前体之间是保守的：章鱼肽NFDEIDRSGFGFN（3个拷贝）、NFDEIDRSGFGFH（2个拷贝）和NFDEIDRSGFGFV（2个拷贝）、FDAFTTGFGHN（一种与促肌动蛋白相关的肽）、SSEDMDRLGFGFN、GDY((SO3))DVYPE、VYGPRDIANLY和SAE。此外，从每种前激素原中预测出两种较长肽中的一种（GPIKVRFLSAIFIPIAAPARSSPQQDAAAGYTDGAPV或APARSSPQQDAAAGYTDGAPV）。基质辅助激光解吸电离傅里叶变换质谱分析证实，在美洲龙虾神经组织中存在所有预测的章鱼肽、与促肌动蛋白相关的肽以及三种与前体相关的肽，即SSEDMDRLGFGFN、GDYDVYPE（未硫酸化）和VYGPRDIANLY。未检测到SAE和较长的、不共享的肽。从克氏原螯虾转录本中预测出类似的肽互补序列；其中大多数通过质谱在其神经组织中被检测到。在这两个物种中还检测到了未从任何前体预测出的截短章鱼肽。与之前对溪蟹的研究一致，NFDEIDRSGFGFN增加了克氏原螯虾中肠/后肠的运动。令人惊讶的是，尽管美洲龙虾中天然存在相同的肽，但它对该物种的肠道运动没有影响。总之，我们的结果为未来研究龙虾类动物中章鱼肽/章鱼肽前体相关肽的调节和生理功能提供了框架。
