Department of Basic Sciences and Social Science for Nursing, Faculty of Nursing, University of Colombo, Thalapathpitiya, 10250, Nugegoda, Sri Lanka.
Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, 102 Jejudaehakno, Ara-Dong, Jeju, Jeju Self-Governing Province, 63243, Republic of Korea.
Mol Biol Rep. 2021 May;48(5):4933-4942. doi: 10.1007/s11033-021-06415-9. Epub 2021 May 26.
Cystatins are reversible inhibitors of cysteine proteases which show an omnipresent distribution in the life on earth. Although, cystatins with mammalian origin were well characterized and their roles in physiology were reported in details, those from teleostean origin are still underrepresented in literature. However, role of cystatins in fish physiology and immune defense is highlighted in few recent reports. In this study, a cystatin C holmologue from rock bream (Oplegnathus fasciatus); termed RbCytC was identified and molecularly characterized. The complete coding sequence of RbCytC was 387 bp in length, which codes for a polypeptide with 129 amino acids, including a signal peptide of 19 amino acids. The consensus cystatin family signatures including a G residue, turning up towards the N-terminus region, QVVAG motif, locating at the middle of the sequence and the PW motif at the c terminal region was found to be well conserved in RbCytC. Phylogenetic analysis using different cystatin counterparts affirmed the close evolutionary relationship of RbCytC with its teleostan homologs which belong to family 2 cystatins. The predicted molecular model of RbCytC resembled most of the structural features of empirically elucidated tertiary structures for chicken egg white cystatin. According to the qPCR assays, RbCytC showed detectable expression in all fish tissues used in the experiment, with markedly pronounced expression level in liver. Moreover, its basal mRNA expression was up-regulated in liver and spleen tissues by experimental rock bream iridovirus infection, whereas down regulated in the same tissues, post live Edwardsiella tarda injection. Collectively, outcomes of our study validate the structural homology of RbCytC with known cystatin C similitudes, especially those of teleosts and suggest its potential roles in proteolytic processes of rock bream physiology as well as in host immune defense mechanisms.
组织蛋白酶抑制剂是半胱氨酸蛋白酶的可逆抑制剂，在地球上的生命中无处不在。尽管具有哺乳动物起源的组织蛋白酶已得到很好的表征，并且其在生理学中的作用已详细报道，但来自硬骨鱼类的组织蛋白酶在文献中仍然代表性不足。然而，最近的一些报道强调了组织蛋白酶在鱼类生理学和免疫防御中的作用。在这项研究中，从石斑鱼（Oplegnathus fasciatus）中鉴定并分子表征了一种组织蛋白酶 C 同系物；称为 RbCytC。RbCytC 的完整编码序列长 387bp，编码 129 个氨基酸的多肽，包括 19 个氨基酸的信号肽。在 RbCytC 中发现了保守的组织蛋白酶家族特征，包括 G 残基，向 N 末端区域扭转，位于序列中部的 QVVAG 基序以及 C 末端区域的 PW 基序。使用不同的组织蛋白酶对应物进行的系统发育分析证实了 RbCytC 与其属于 2 型组织蛋白酶家族的硬骨鱼类同源物的密切进化关系。RbCytC 的预测分子模型与经验上阐明的鸡卵清白蛋白组织蛋白酶的三级结构的大多数结构特征相似。根据 qPCR 测定，RbCytC 在实验中使用的所有鱼类组织中均有可检测的表达，在肝脏中表达水平明显较高。此外，在实验性石斑鱼虹彩病毒感染后，其肝脏和脾脏组织中的基础 mRNA 表达上调，而在相同组织中，在注射爱德华氏菌后表达下调。总之，我们的研究结果验证了 RbCytC 与已知组织蛋白酶 C 相似物的结构同源性，尤其是硬骨鱼类的相似性，并表明其在石斑鱼生理学的蛋白水解过程以及宿主免疫防御机制中具有潜在作用。
