The majority of mouse and human genes are subject to alternative cleavage and polyadenylation (APA), which most often leads to the expression of two or more alternative length 3' untranslated region (3' UTR) mRNA isoforms. In neural tissues, there is enhanced expression of APA isoforms with longer 3' UTRs on a global scale, but the physiological relevance of these alternative 3' UTR isoforms is poorly understood. Calmodulin 1 (Calm1) is a key integrator of calcium signaling that generates short (Calm1-S) and long (Calm1-L) 3' UTR mRNA isoforms via APA. We found Calm1-L expression to be largely restricted to neural tissues in mice including the dorsal root ganglion (DRG) and hippocampus, whereas Calm1-S was more broadly expressed. smFISH revealed that both Calm1-S and Calm1-L were subcellularly localized to neural processes of primary hippocampal neurons. In contrast, cultured DRG showed restriction of Calm1-L to soma. To investigate the in vivo functions of Calm1-L, we implemented a CRISPR-Cas9 gene editing strategy to delete a small region encompassing the Calm1 distal polyA site. This eliminated Calm1-L expression while maintaining expression of Calm1-S. Mice lacking Calm1-L (Calm1ΔL/ΔL) exhibited disorganized DRG migration in embryos, and reduced experience-induced neuronal activation in the adult hippocampus. These data indicate that Calm1-L plays functional roles in the central and peripheral nervous systems.

中文翻译：

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通过 CRISPR-Cas9 基因编辑消除 Calm1 长 3' UTR mRNA 异构体损害小鼠背根神经节发育和海马神经元激活

大多数小鼠和人类基因都会发生选择性切割和多聚腺苷酸化 (APA)，这通常会导致表达两个或多个长度不同的 3' 非翻译区 (3' UTR) mRNA 同种型。在神经组织中，在全球范围内具有更长 3' UTR 的 APA 同种型的表达增强，但这些替代 3' UTR 同种型的生理相关性知之甚少。钙调蛋白 1 (Calm1) 是钙信号传导的关键整合者，可通过 APA 产生短 (Calm1-S) 和长 (Calm1-L) 3' UTR mRNA 异构体。我们发现 Calm1-L 的表达主要限于小鼠的神经组织，包括背根神经节 (DRG) 和海马，而 Calm1-S 的表达更广泛。smFISH 显示 Calm1-S 和 Calm1-L 均亚细胞定位于原代海马神经元的神经过程。相比之下，培养的 DRG 显示 Calm1-L 对体细胞的限制。为了研究 Calm1-L 的体内功能，我们实施了 CRISPR-Cas9 基因编辑策略来删除包含 Calm1 远端 polyA 位点的小区域。这消除了 Calm1-L 的表达，同时保持 Calm1-S 的表达。缺乏 Calm1-L (Calm1ΔL/ΔL) 的小鼠在胚胎中表现出无序的背根神经节迁移，并减少了成年海马中经验诱导的神经元激活。这些数据表明 Calm1-L 在中枢和外周神经系统中发挥功能作用。我们实施了 CRISPR-Cas9 基因编辑策略来删除包含 Calm1 远端 polyA 位点的小区域。这消除了 Calm1-L 的表达，同时保持 Calm1-S 的表达。缺乏 Calm1-L (Calm1ΔL/ΔL) 的小鼠在胚胎中表现出无序的背根神经节迁移，并减少了成年海马中经验诱导的神经元激活。这些数据表明 Calm1-L 在中枢和外周神经系统中发挥功能作用。我们实施了 CRISPR-Cas9 基因编辑策略来删除包含 Calm1 远端 polyA 位点的小区域。这消除了 Calm1-L 的表达，同时保持 Calm1-S 的表达。缺乏 Calm1-L (Calm1ΔL/ΔL) 的小鼠在胚胎中表现出无序的背根神经节迁移，并减少了成年海马中经验诱导的神经元激活。这些数据表明 Calm1-L 在中枢和外周神经系统中发挥功能作用。