The role of miRNAs and lncRNAs in regulating transcription in gilthead sea bream (Sparus aurata) myoblasts in response to amino acids and insulin-like growth factor 1 [RNA-seq]

In this study, gilthead sea bream (Sparus aurata) fast muscle myoblasts were stimulated with two pro-growth treatments, amino acids (AA) and insulin-like growth factor 1 (Igf-1), to analyze the transcriptional response of genes, microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) and their regulatory network. AA had a higher impact on gene transcription (1795 genes significantly changed) compared to Igf-1 (385 genes significantly changed). Both treatments stim-ulated the transcription of genes related to muscle differentiation (GO:0042692) and sarcomere components (GO:0030017), but AA stimulated more the DNA replication and cell division (GO:0007049). Notably, four miRNAs (miR-21, miR-146, miR-22b and miR-206) dominated the landscape among 403 expressed miRNAs. Both pro-growth treatments altered the transcription of over 100 miRNAs, including muscle-specific miRNAs (myomiRs) such as miR-133a/b, miR-206, miR-499, miR-1, and miR 27a. Among 111 detected lncRNAs (> 1 FPKM), only 30 were significantly changed by AA and 11 by Igf-1. Eight lncRNAs exhibited strong negative correlations with several mRNAs, suggesting direct regulation; while 30 lncRNAs showed strong correlations and interac-tions with several miRNAs, suggesting their function as miRNA's sponges. This work is the first step in the identification of ncRNAs network controlling muscle development and growth in gilthead sea bream, pointing out potential regulatory mechanisms in response to pro-growth signals. Overall design: To obtain the transcriptional profile of gilthead sea bream (Sparus aurata) muscle cells treated with amino acids (AA), IGF1 or cultured in nutrient restriction medium (Control), with n=3 for each experimental group.

本研究以金头鲷（Sparus aurata）快肌成肌细胞为实验材料，采用两种促生长处理——氨基酸（Amino acids, AA）与胰岛素样生长因子1（Insulin-like growth factor 1, Igf-1）进行刺激，旨在分析基因、微小RNA（microRNAs, miRNAs）及长链非编码RNA（long non-coding RNAs, lncRNAs）的转录响应及其调控网络。相较于Igf-1处理组（仅385个基因发生显著表达变化），氨基酸处理对基因转录的影响更为显著，共涉及1795个差异表达基因。两种处理均能激活与肌肉分化（GO:0042692）及肌节组分（GO:0030017）相关的基因转录，但氨基酸处理更显著地促进了DNA复制与细胞分裂（GO:0007049）相关通路的激活。值得注意的是，在403个已检测到表达的miRNAs中，miR-21、miR-146、miR-22b及miR-206这4种miRNA占据表达主导地位。两种促生长处理均能改变超过100种miRNAs的转录水平，其中包括肌特异性miRNAs（肌源miRNAs, myomiRs）如miR-133a/b、miR-206、miR-499、miR-1及miR-27a。在111个检测到的lncRNAs（FPKM>1）中，仅30个经氨基酸处理后发生显著表达变化，Igf-1处理组则仅11个。其中8个lncRNAs与多个mRNA呈现强烈负相关，提示其可能直接调控mRNA表达；另有30个lncRNAs与多种miRNAs存在强相关性及相互作用，表明其可能作为miRNA海绵发挥功能。本研究是鉴定金头鲷肌肉发育与生长相关非编码RNA调控网络的第一步，阐明了其响应促生长信号的潜在调控机制。实验设计概况：本研究旨在获取经氨基酸（AA）、IGF1处理，或于营养限制培养基中培养的金头鲷肌肉细胞的转录组谱，每组设置3个生物学重复。