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一种新型 Cre 重组酶小鼠品系,用于在形成 ribbon 突触的视网膜神经元中特异性删除 floxed 基因。

A Novel Cre Recombinase Mouse Strain for Cell-Specific Deletion of Floxed Genes in Ribbon Synapse-Forming Retinal Neurons.

机构信息

Institute of Anatomy and Cell Biology, Department of Neuroanatomy, Medical School, Saarland University, 66421 Homburg, Germany.

Institute of Clinical and Experimental Pharmacology, Center for Molecular Signaling (PZMS) and Center for Gender-Specific Biology and Medicine (CGBM), Medical School, Saarland University, 66421 Homburg, Germany.

出版信息

Int J Mol Sci. 2024 Feb 5;25(3):1916. doi: 10.3390/ijms25031916.

Abstract

We generated a novel Cre mouse strain for cell-specific deletion of floxed genes in ribbon synapse-forming retinal neurons. Previous studies have shown that the RIBEYE promotor targets the expression of recombinant proteins such as fluorescently tagged RIBEYE to photoreceptors and retinal bipolar cells and generates fluorescent synaptic ribbons in situ in these neurons. Here, we used the same promotor to generate a novel transgenic mouse strain in which the RIBEYE promotor controls the expression of a Cre-ER(T2) recombinase (RIBEYE-Cre). To visualize Cre expression, the RIBEYE-Cre animals were crossed with ROSA26 tau-GFP (R26-τGFP) reporter mice. In the resulting RIBEYE-Cre/R26 τGFP animals, Cre-mediated removal of a transcriptional STOP cassette results in the expression of green fluorescent tau protein (tau-GFP) that binds to cellular microtubules. We detected robust tau-GFP expression in retinal bipolar cells. Surprisingly, we did not find fluorescent tau-GFP expression in mouse photoreceptors. The lack of tau-GFP reporter protein in these cells could be based on the previously reported absence of tau protein in mouse photoreceptors which could lead to the degradation of the recombinant tau protein. Consistent with this, we detected Cre and tau-GFP mRNA in mouse photoreceptor slices by RT-PCR. The transgenic RIBEYE-Cre mouse strain provides a new tool to study the deletion of floxed genes in ribbon synapse-forming neurons of the retina and will also allow for analyzing gene deletions that are lethal if globally deleted in neurons.

摘要

我们构建了一种新型 Cre 小鼠品系,用于在形成神经纤维层突触的视网膜神经元中特异性敲除 floxed 基因。先前的研究表明,RIBEYE 启动子可将重组蛋白(如荧光标记的 RIBEYE)的表达靶向至光感受器和视网膜双极细胞,并在这些神经元中体内产生荧光性神经纤维层突触。在这里,我们使用相同的启动子构建了一种新型转基因小鼠品系,其中 RIBEYE 启动子控制 Cre-ER(T2) 重组酶(RIBEYE-Cre)的表达。为了可视化 Cre 表达,将 RIBEYE-Cre 动物与 ROSA26tau-GFP(R26-τGFP)报告小鼠进行杂交。在产生的 RIBEYE-Cre/R26-τGFP 动物中,Cre 介导的转录终止盒的去除导致绿色荧光 tau 蛋白(tau-GFP)的表达,该 tau-GFP 与细胞微管结合。我们在视网膜双极细胞中检测到了强烈的 tau-GFP 表达。令人惊讶的是,我们在小鼠光感受器中未发现荧光性 tau-GFP 表达。这些细胞中缺乏 tau-GFP 报告蛋白可能基于先前报道的小鼠光感受器中缺乏 tau 蛋白,这可能导致重组 tau 蛋白的降解。与此一致的是,我们通过 RT-PCR 在小鼠光感受器切片中检测到 Cre 和 tau-GFP mRNA。该转基因 RIBEYE-Cre 小鼠品系为研究视网膜形成神经纤维层突触的神经元中 floxed 基因的缺失提供了一种新工具,并且还将允许分析如果在神经元中全局缺失则具有致死性的基因缺失。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5f3d/10856425/dbf150551e80/ijms-25-01916-g005.jpg

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