在早期胚胎发育中,转录因子和信号通路的交叉调节在介导发育和生理过程中至关重要。此外,许多研究表明,microRNAs(miRNAs)介导的信号和网络成分的转录后调控的重要性;然而,miRNAs是如何被转录调控的,人们知之甚少。miRNAs是许多生物过程的关键微调因子,它们的失调会导致多种疾病和发育缺陷。以前,我们已经证明miRNAs在整个海胆发育过程中动态表达,这表明miRNAs可能处于转录调控之下。这里,我们使用了药物抑制剂,遗传结构,和功能丧失试剂,以评估关键信号通路的影响(Wnt,节点,MAPK,刺猬索尼克,三角洲/缺口,VEGF,和BMP)和转录因子(Alx1,Ets1/2和Tbr)在进化保守的miR-1,miR-31,miR-92和miR-124的转录水平上;无脊椎动物特异性miR-71;和棘皮动物特异性miR-2002,miR-2007和miR-2012。我们还使用计算方法来鉴定这些miRNA的潜在转录因子结合位点。从MEME-SuiteMotif数据库获得转录因子(TF)的结合基序列表,并用作算法FIMO(查找个体基序发生)的输入,它在较大的序列中检测短的核苷酸基序。基于miRNA表达的实验数据结合生物信息学预测,我们认为转录因子Tbr,Alx1和Ets1分别调节SpmiR-1、SpmiR-31和SpmiR-71。我们还观察到由于对Wnt的扰动对miRNA水平的显著影响,节点,MAPK,和SonicHedgehog信号通路,虽然在对Delta/Notch的扰动下没有观察到miRNA水平的显著变化,VEGF,或BMP信号通路。总的来说,这项研究提供了通过信号通路和转录因子对miRNAs转录调控的见解,并有助于我们对发育过程的遗传调控的整体理解。
In early embryonic development, the cross-regulation of transcription factors and signaling pathways are critical in mediating developmental and physiological processes. Additionally, many studies have shown the importance of post-transcriptional regulation of signaling and network components mediated by microRNAs (miRNAs); however, how miRNAs are transcriptionally regulated is poorly understood. miRNAs are critical fine-tuners of many biological processes and their dysregulation leads to a variety of diseases and developmental defects. Previously, we have shown that miRNAs are dynamically expressed throughout sea urchin development, suggesting that miRNAs are likely to be under transcriptional regulation. Here, we used pharmacological inhibitors, genetic constructs, and loss-of-function reagents to assess the impact of key signaling pathways (Wnt, Nodal, MAPK, Sonic Hedgehog, Delta/Notch, VEGF, and BMP) and transcription factors (Alx1, Ets1/2, and Tbr) on the transcript levels of the evolutionarily conserved miR-1, miR-31, miR-92 and miR-124; the invertebrate-specific miR-71; and the echinoderm-specific miR-2002, miR-2007, and miR-2012. We also used computational methods to identify potential transcription factor binding sites of these miRNAs. Lists of binding motifs for transcription factors (TFs) were acquired from the MEME-Suite Motif Database and used as inputs for the algorithm FIMO (Find Individual Motif Occurrences), which detects short nucleotide motifs within larger sequences. Based on experimental data on miRNA expression in conjunction with bioinformatic predictions, we propose that the transcription factors Tbr, Alx1, and Ets1 regulate SpmiR-1, SpmiR-31, and SpmiR-71, respectively. We additionally observed significant effects on miRNA levels as a result of perturbations to Wnt, Nodal, MAPK, and Sonic Hedgehog signaling pathways, while no significant change on miRNA levels were observed with perturbations to Delta/Notch, VEGF, or BMP signaling pathways. Overall, this study provides insights into the transcriptional regulation of miRNAs by signaling pathways and transcription factors and contribute to our overall understanding of the genetic regulation of developmental processes.