Laboratory of Guangdong Province and Hong Kong Region on Marine Bioresource Conservation and Exploitation, College of Marine Sciences, South China Agricultural University, Guangzhou, 510642, China; Guangdong Modern Agricultural (Quality and Safety of Aquatic Products) Industrial Technology R&D Center, Guangzhou, 510642, China.
Laboratory of Guangdong Province and Hong Kong Region on Marine Bioresource Conservation and Exploitation, College of Marine Sciences, South China Agricultural University, Guangzhou, 510642, China.
Fish Shellfish Immunol. 2022 Aug;127:918-924. doi: 10.1016/j.fsi.2022.07.021. Epub 2022 Jul 19.
PAP (3'-phosphoadenosine 5'-phosphate) is a ubiquitous phosphoric acid and a natural inhibitor of the XRN (5'-3'exoribonuclease) family. It was proved to enter the nucleus through the retrograde signaling pathway and inhibit XRN2 to prevent the degradation of miRNA precursors, thus promoting the anti-oxidation miRNA level in Arabidopsis thaliana. Vitamin E (tocopherol) was proved to promote the accumulation of PAP in the plant, which facilitates PAP into the nucleus to accomplish its antioxidant function. However, the relationship between VE and PAP in animals is unclear. To identify the relationship between VE and PAP and to uncover the function of PAP in fish, we investigated the performance of VE and PAP in Nile tilapia by comparing the antioxidant indicators (SOD, GSH-Px, and CAT), the Keap1-Nrf2 signaling pathway, and the miRNA expression profiles. Results showed that the antioxidant effect of VE and PAP showed similar character either in tilapia liver or in serum: the activities of GSH-Px and CAT of both groups were significantly increased (P < 0.05); the SOD activity of the VE group was significantly increased (P < 0.05), and although the result of the PAP group was not so significant (P > 0.05), PAP improved the SOD level, too. The two groups also showed similar character in the tilapia liver; both did not significantly increase the liver δ-VE content (P > 0.05). However, VE significantly increased the content of α-VE and γ-VE (P < 0.05), while the PAP group was insignificant (P > 0.05). Feed with VE and intraperitoneal injection of PAPs reagent both increased the PAP content in the liver of tilapia, and the effect of the VE group was more significant (P < 0.05) than that of the PAP group (P > 0.05). Both groups reduced the expression of Keap1 and Cullin3 genes and improved the level of HO-1 gene expression, with the improved miRNA level of Nrf2. As a logical result, they decreased the expression of XRN1 and XRN2. By profile sequencing, we further identified some antioxidant closely related miRNAs shared in the VE and PAP groups, including miR-30, miR-24, miR-19b, and miR-100. By comparing the regulating mechanism of VE and PAP of feed supply and intraperitoneal injection, we proved that VE and PAP were closely related in fish; VE promoted the gathering of PAP. The latter retrograded into the nucleus of the fish liver to inhibit the expression of XRN genes and to up-regulate antioxidant miRNA levels as it does in plants. Only the PAP can accomplish the antioxidant activities, while VE promotes the process. Our study laid the foundation for the application of PAP as a new antioxidant agent in fish farming and benefit a further understanding of the VE antioxidant function in fish.
PAP（3'-磷酸腺苷 5'-磷酸）是一种普遍存在的磷酸和天然的 XRN（5'-3'外切核酸酶）家族抑制剂。已证明它通过逆行信号通路进入细胞核，抑制 XRN2 以防止 miRNA 前体降解，从而促进拟南芥中的抗氧化 miRNA 水平。维生素 E（生育酚）被证明可促进植物中 PAP 的积累，从而有利于 PAP 进入细胞核以完成其抗氧化功能。然而，VE 和 PAP 在动物中的关系尚不清楚。为了确定 VE 和 PAP 之间的关系并揭示 PAP 在鱼类中的功能，我们通过比较抗氧化指标（SOD、GSH-Px 和 CAT）、Keap1-Nrf2 信号通路和 miRNA 表达谱，研究了尼罗罗非鱼中 VE 和 PAP 的表现。结果表明，VE 和 PAP 的抗氧化作用在罗非鱼肝或血清中表现出相似的特征：两组的 GSH-Px 和 CAT 活性均显著增加（P<0.05）；VE 组的 SOD 活性显著增加（P<0.05），尽管 PAP 组的结果不那么显著（P>0.05），但 PAP 也提高了 SOD 水平。两组在罗非鱼肝中也表现出相似的特征；均未显著增加肝 δ-VE 含量（P>0.05）。然而，VE 显著增加了 α-VE 和 γ-VE 的含量（P<0.05），而 PAP 组则不显著（P>0.05）。VE 喂养和 PAP 试剂腹腔注射均增加了罗非鱼肝中的 PAP 含量，VE 组的效果比 PAP 组更显著（P<0.05）。两组均降低了 Keap1 和 Cullin3 基因的表达，并提高了 HO-1 基因的表达水平，同时提高了 Nrf2 的 miRNA 水平。作为合乎逻辑的结果，它们降低了 XRN1 和 XRN2 的表达。通过基因表达谱测序，我们进一步鉴定了 VE 和 PAP 组中一些与抗氧化密切相关的 miRNA，包括 miR-30、miR-24、miR-19b 和 miR-100。通过比较饲料供应和腹腔注射的 VE 和 PAP 的调节机制，我们证明了 VE 和 PAP 在鱼类中密切相关；VE 促进了 PAP 的聚集。后者逆行进入鱼肝细胞核，抑制 XRN 基因的表达，并上调抗氧化 miRNA 水平，就像在植物中一样。只有 PAP 才能完成抗氧化活性，而 VE 则促进这一过程。我们的研究为 PAP 作为一种新型抗氧化剂在水产养殖中的应用奠定了基础，并有助于进一步了解 VE 在鱼类中的抗氧化功能。
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