Molecular, mass spectral, and physiological analyses of orcokinins and orcokinin precursor-related peptides in the lobster Homarus americanus and the crayfish Procambarus clarkii.
Molecular, mass spectral, and physiological analyses of orcokinins and orcokinin precursor-related peptides in the lobster Homarus americanus and the crayfish Procambarus clarkii.
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DOI：
10.1016/j.peptides.2008.10.009
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发表时间：
2009-03
期刊：
影响因子：
3
通讯作者：
Christie AE
中科院分区：
文献类型：
--
作者：
Dickinson PS;Stemmler EA;Barton EE;Cashman CR;Gardner NP;Rus S;Brennan HR;McClintock TS;Christie AE
最近，从克氏原螯虾中克隆了编码前原-orcokinins 的 cDNA。这些 cDNA 编码四种 orcokinin 异构体以及几种其他肽的多个拷贝。使用克氏虾转录本的翻译开放阅读框作为查询，通过 BLAST 分析鉴定出编码美洲龙虾 Homarus americanus 直向同源物的 5 个 EST。从这些克隆中，鉴定出三种 cDNA，每种编码两种不同的前激素原之一。推导的前激素原的预测加工将产生 13 个肽，其中 12 个在两个前体之间保守：促激肽 NFDEIDRSGFGFN（3 个拷贝）、NFDEIDRSGFGFH（2 个拷贝）和 NFDEIDRSGFGFV（2 个拷贝）、FDAFTTGFGHN（促促肌肽相关肽）、SSEDMDRLGFGFN、GDY(SO3)DVYPE、 VYGPRDIANLY 和 SAE。此外，从每个前激素原预测两个较长肽之一（GPIKVRFLSAIFIPIAAPARSSPQQDAAAGYTDGAPV、APARSSPQQDAAAGYTDGAPV）。 MALDI-FTMS 分析证实，美洲人神经组织中存在所有预测的 orcokinins、orcomyotropin 相关肽和三种前体相关肽 SSEDMDRLGFGFN、GDYDVYPE（未硫酸化）和 VYGPRDIANLY。未检测到 SAE 和较长的非共享肽。从克氏伯克酵母转录物中预测出类似的肽互补体；其中大部分是通过质谱在其神经组织中检测到的。在这两个物种中也检测到了未从任何前体预测到的截短的 orcokinins。与之前对小龙虾 Orconectes limosus 的研究一致，NFDEIDRSGFGFN 增加了克氏对虾的中肠/后肠运动。令人惊讶的是，相同的肽虽然是美洲人天然的，但并不影响该物种的肠道蠕动。总之，我们的结果为未来研究虾类中 orcokinins/orcokinin 前体相关肽的调节和生理功能提供了框架。
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 two 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, 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.
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