College of Ecology, Lishui University, Lishui 323000, China.
College of Ecology, Lishui University, Lishui 323000, China.
Comp Biochem Physiol C Toxicol Pharmacol. 2021 Sep;247:109072. doi: 10.1016/j.cbpc.2021.109072. Epub 2021 May 7.
Cathelicidins are an important antimicrobial peptide family and are expressed in many different vertebrates. They play an important role in the innate immune system of the host. However, amphibian cathelicidins are poorly understood. In this study, the cDNA of the cathelicidin gene was obtained from the skin transcriptome of tiger frog (Hoplobatrachus rugulosus). The predicted amino acid sequence of tiger frog cathelicidin (HR-CATH) comprises a signal peptide, a cathelin domain, and a mature peptide. The HR-CATH amino acid sequence alignment with other frog cathelicidins showed that the functional mature peptide is highly variable in amphibians, whereas the cathelin domain is conserved. A phylogenetic tree analysis showed that HR-CATH is most closely related to cathelicidin-NV from Nanorana ventripunctata. HR-CATH was chemically synthesized and its in vitro activity was determined. It had high antibacterial activity against Vibrio parahaemolyticus, Staphylococcus aureus, and the pathogenic bacterium Aeromonas hydrophila. HR-CATH damaged the cell membrane integrity of A. hydrophila according to a lactate dehydrogenase release assay and was able to hydrolyze the genomic DNA from A. hydrophila in a dose-dependent manner. Furthermore, in RAW264.7 cells (mouse leukemic monocyte/macrophage cell line), HR-CATH induced chemotaxis and enhanced respiratory burst. Our study shows that amphibian cathelicidin has antimicrobial activity and an immunomodulatory effect on immune cells.
抗菌肽是一类重要的抗菌肽家族，存在于许多不同的脊椎动物中。它们在宿主的先天免疫系统中发挥着重要作用。然而，关于两栖动物抗菌肽的研究还知之甚少。本研究从虎纹蛙（Hoplobatrachus rugulosus）皮肤转录组中获得了抗菌肽基因的 cDNA。虎纹蛙抗菌肽（HR-CATH）的预测氨基酸序列包含一个信号肽、一个抗菌肽前体和一个成熟肽。与其他蛙类抗菌肽的氨基酸序列比对表明，功能成熟肽在两栖动物中高度多变，而抗菌肽前体域则保守。系统进化树分析表明，HR-CATH 与 Nanorana ventripunctata 的抗菌肽-NV 最为密切相关。HR-CATH 被化学合成，并测定了其体外活性。它对副溶血弧菌、金黄色葡萄球菌和致病性嗜水气单胞菌具有很高的抗菌活性。根据乳酸脱氢酶释放试验，HR-CATH 破坏了嗜水气单胞菌的细胞膜完整性，并能以剂量依赖的方式水解嗜水气单胞菌的基因组 DNA。此外，在 RAW264.7 细胞（小鼠白血病单核/巨噬细胞系）中，HR-CATH 诱导趋化作用并增强呼吸爆发。本研究表明，两栖动物抗菌肽具有抗菌活性，并对免疫细胞具有免疫调节作用。
