Department of Ophthalmology and Visual Sciences, University of Wisconsin-Madison, Madison, WI, 53706, USA.
Department of Ophthalmology and Visual Sciences, University of Wisconsin-Madison, Madison, WI, 53706, USA; Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, WI, 53706, USA; McPherson Eye Research Institute, University of Wisconsin-Madison, Madison, WI, 53706, USA.
Exp Eye Res. 2023 Nov;236:109647. doi: 10.1016/j.exer.2023.109647. Epub 2023 Sep 7.
The retinoic acid-inducible gene I (RIG)-I-like receptor (RLR) family of RNA sensor proteins plays a key role in the innate immune response to viral nucleic acids, including viral gene delivery vectors, but little is known about the expression of RLR proteins in the retina. The purpose of this study was to characterize cell-specific expression patterns of RLR proteins in non-human primate (NHP) neural retina tissue and to examine if RLR pathway signaling restricts viral gene delivery transduction. Since RLR protein signaling converges at the mitochondrial antiviral signaling protein (MAVS), experiments were performed to determine if knockdown of MAVS affected FIVGFP transduction efficiency in the human Mueller cell line MIO-M1. Immunoblotting confirmed expression of RIG-I, melanoma differentiation-associated protein 5 (MDA5), laboratory of genetics and physiology 2 (LGP2), and MAVS proteins in MIO-M1 cells and NHP retina tissue. Double label immunofluorescence (IF) studies revealed RIG-I, LGP2, and MAVS were expressed in Mueller microglial cells in the NHP retina. In addition, LGP2 and MDA5 proteins were detected in cone and retinal ganglion cells (RGC). MDA5 was also present in a subset of calretinin positive amacrine cells, and in nuclei within the inner nuclear layer (INL). Knockdown of MAVS significantly increased the transduction efficiency of the lentiviral vector FIVGFP in MIO-M1 cells, compared to control cells. FIVGFP or AAVGFP challenge did not alter expression of the LGP2, MAVS, MDA5 or RIG-I genes in MIO-M1 cells or NHP retina tissue compared to media treated controls. Our data demonstrate that innate immune response proteins involved in viral RNA sensing, including MDA5, RIG-I, LGP2, and MAVS, are expressed in several cell types within the NHP neural retina. In addition, the MAVS protein restricts non-human lentiviral transduction efficiency in MIO-M1 cells.
视黄酸诱导基因 I（RIG）-I 样受体（RLR）家族的 RNA 传感器蛋白在对病毒核酸的先天免疫反应中发挥着关键作用，包括病毒基因传递载体，但对于 RLR 蛋白在视网膜中的表达知之甚少。本研究的目的是描述非人类灵长类动物（NHP）神经视网膜组织中 RLR 蛋白的细胞特异性表达模式，并研究 RLR 通路信号是否限制病毒基因传递转导。由于 RLR 蛋白信号在抗病毒信号蛋白（MAVS）处汇聚，因此进行了实验以确定 MAVS 的敲低是否会影响 FIVGFP 在人 Mueller 细胞系 MIO-M1 中的转导效率。免疫印迹证实了 RIG-I、黑色素瘤分化相关蛋白 5（MDA5）、遗传学和生理学实验室 2（LGP2）和 MAVS 蛋白在 MIO-M1 细胞和 NHP 视网膜组织中的表达。双标免疫荧光（IF）研究显示 RIG-I、LGP2 和 MAVS 在 NHP 视网膜中的 Mueller 小胶质细胞中表达。此外，LGP2 和 MDA5 蛋白在视锥细胞和视网膜神经节细胞（RGC）中检测到。MDA5 也存在于一部分 calretinin 阳性无长突细胞和内核层（INL）内的核中。与对照细胞相比，MAVS 的敲低显著增加了 MIO-M1 细胞中慢病毒载体 FIVGFP 的转导效率。与对照相比，FIVGFP 或 AAVGFP 挑战并未改变 MIO-M1 细胞或 NHP 视网膜组织中 LGP2、MAVS、MDA5 或 RIG-I 基因的表达。我们的数据表明，参与病毒 RNA 感应的先天免疫反应蛋白，包括 MDA5、RIG-I、LGP2 和 MAVS，在 NHP 神经视网膜中的几种细胞类型中表达。此外，MAVS 蛋白限制了 MIO-M1 细胞中非人类慢病毒的转导效率。
