Transcriptome analysis of the post-larvae of giant freshwater prawn (Macrobrachium rosenbergii) after IAG gene knockdown with microRNA interference
Transcriptome analysis of the post-larvae of giant freshwater prawn (Macrobrachium rosenbergii) after IAG gene knockdown with microRNA interference
复制标题
通过 microRNA 干扰敲除 IAG 基因后罗氏沼虾幼体的转录组分析
DOI：
10.1016/j.ygcen.2022.114054
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发表时间：
2022-05-17
影响因子：
2.7
通讯作者：
Qiu, Gaofeng
中科院分区：
文献类型：
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作者：
Qian, Hongli;Ma, Keyi;Qiu, Gaofeng
甲壳类动物的类胰岛素雄激素基因（IAG）在性别分化的调节中发挥着关键作用。 MicroRNA (miRNA) 是一类短的非编码 RNA，充当转录后基因调节因子。然而，有关miRNA与罗氏沼虾IAG（MrIAG）之间调控关系的信息却很少。在本研究中，我们使用 MrIAG 的 3' 非翻译区 (UTR) 来预测 miRNA 的潜在靶位点。结果表明，miR-184在MrIAG的3'UTR上有1个靶位点。体外双荧光素酶报告测定证实miR-184可以显着下调MrIAG的表达。此外，我们构建了MrIAG 3'UTR的突变质粒。结果显示，突变质粒与miR-184 agomir共转染后，荧光素酶活性与对照相比并未受到影响。这些结果表明miR-184可以直接调节MrIAG。此外，我们发现罗氏沼虾中miR-184的过度表达可导致基因转录水平的显着变化。与对照组相比，我们在miR-184注射组中鉴定出1510个差异表达基因（DEG）。一些DEGs被发现参与性别分化、性腺发育、生长和蜕皮。随机对8个DEG进行qRT-PCR验证，结果显示，MrIAG基因敲除后，性别、生长、代谢相关基因的表达水平发生显着变化。总的来说，这项研究的结果表明，miR-184 通过介导 IAG 表达，可能参与罗氏沼虾的许多生理过程。本研究为miR-184短期沉默MrIAG奠定了基础，为今后罗氏沼虾的miRNA功能分析提供了基础。
The insulin-like androgenic gland hormone gene (IAG) of crustaceans plays pivotal roles in the regulation of sex differentiation. MicroRNAs (miRNAs) are a class of short, non-coding RNAs that function as post-transcriptional gene regulators. However, little information about the regulatory relationship between miRNA and Macro-brachium rosenbergii IAG (MrIAG) were exposed. In this study, we used the 3' untranslated region (UTR) of MrIAG to predict potential target sites of miRNAs. The results showed that miR-184 has one target site in the 3' UTR of MrIAG. Dual-luciferase report assay in vitro confirmed that miR-184 can significantly down-regulate MrIAG expression. Besides, we constructed mutant plasmids of 3' UTR of MrIAG. The result displayed that after co-transfection of mutant plasmids and miR-184 agomir, the activity of luciferase was not affected compared to the control. These results indicated that miR-184 could directly regulate MrIAG. In addition, we found that overexpression of miR-184 in M. rosenbergii can lead to significant changes in the transcription level of genes. Compared with control group, we identified 1510 differentially expressed genes (DEGs) in the miR-184 injection group. Some DEGs were involved in sex differentiation, gonad development, growth and molting were found. qRT-PCR verification was performed on eight DEGs randomly, and the results showed that the expression level of sex-, growth-, and metabolism-related genes changed significantly after MrIAG gene knockdown. Collectively, findings from this study suggest that miR-184, by mediating IAG expression, may be involved in many physio-logical processes in M. rosenbergii. The current study lays a basic understanding for short-term silencing of MrIAG with miR-184, and facilitates miRNA function analysis in M. rosenbergii in future.