Identification of Abundant and Evolutionarily Conserved Opioid Receptor Circular RNAs in the Nervous System Modulated by Morphine
Identification of Abundant and Evolutionarily Conserved Opioid Receptor Circular RNAs in the Nervous System Modulated by Morphine
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DOI：
10.1124/mol.118.113977
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发表时间：
2019-08-01
影响因子：
3.6
通讯作者：
Pan, Ying-Xian
中科院分区：
文献类型：
--
作者：
He, Takeshi;Shum, Rebecca;Pan, Ying-Xian
环状rna （circRNAs）是通过反剪接形成的单链共价封闭rna的一个独特类别。circrna的功能尚不完全清楚，目前正在积极研究中。在这里，我们报道，除了传统的线性mrna (linRNA)，小鼠，大鼠和人类阿片受体基因产生外显子环状rna亚型。使用标准分子生物学方法，在啮齿动物和人类大脑、脊髓以及人类神经母细胞瘤细胞的rna中检测到Oprm1环状rna (circOprm1)，提示进化守恒。测序证实使用规范剪接位点进行反向剪接。Oprm1环状rna是抗RNase R酶切的义链环状rna。通过定量逆转录聚合酶链反应测定的Oprm1 circRNA与linRNA的相对丰度在小鼠脑区域之间存在差异，circRNA亚型在吻侧结构中占主导地位，在脑干中较少丰度。慢性吗啡暴露小鼠脑circOprm1.e2.3和circOprm1.e2.3增加。e4（302）水平是linRNA的1.5- 1.6倍。序列分析预测在Oprm1环状rna序列中存在大量microRNA结合位点，提示其在海绵吸附microRNA中具有潜在作用。此外，我们观察到其他阿片受体基因，包括delta、kappa和伤害肽受体基因也产生了类似的环状rna。综上所述，阿片受体基因家族的所有成员都表达circRNA，其中Oprm1 circRNA在某些区域的表达水平超过了线性形式。意义声明吗啡对Oprm1环状RNA （circRNA）表达的调节，加上与circRNA序列的潜在miRNA结合位点的高富集度和存在，表明Oprm1环状RNA在慢性阿片样物质效应（如耐受性）中的潜在作用。
Circular RNAs (circRNAs) are a distinct category of single-stranded, covalently closed RNAs formed by backsplicing. The functions of circRNAs are incompletely known and are under active investigation. Here, we report that in addition to traditional linear mRNAs (linRNA), mouse, rat, and human opioid receptor genes generate exonic circRNA isoforms. Using standard molecular biologic methods, Oprm1 circRNAs (circOprm1) were detected in RNAs of rodent and human brains and spinal cords, as well as human neuroblastoma cells, suggesting evolutionary conservation. Sequencing confirmed backsplicing using canonical splice sites. Oprm1 circRNAs were sense-stranded circRNAs resistant to RNase R digestion. The relative abundance of Oprm1 circRNA to linRNA determined by quantitative reverse transcription polymerase chain reaction varied among mouse brain regions, with circRNA isoforms predominating in rostral structures and less abundant in brain stem. Chronic morphine exposure in mice increased brain circOprm1.e2.3 and circOprm1.e2.e3.e4(302) levels by 1.5-to 1.6-fold relative to linRNA. Sequence analysis predicted numerous microRNA binding sites within Oprm1 circRNA sequences, suggesting a potential role in microRNA sequestration through sponging. In addition, we observed that other opioid receptor genes including delta, kappa, and nociceptin receptor genes produced similar circRNAs. In conclusion, all members of the opioid receptor gene family express circRNAs, with Oprm1 circRNA levels exceeding those of linear forms in some regions.SIGNIFICANCE STATEMENTThe modulation of Oprm1 circular RNA (circRNA) expression by morphine, coupled with the high abundance and existence of potential miRNA binding sites with circRNA sequences suggests the potential role of Oprm1 circRNAs in chronic opioid effects such as tolerance.