Department of Pathology, University of California at San Diego, La Jolla, California, USA.
Invest Ophthalmol Vis Sci. 2012 Oct 1;53(11):7159-66. doi: 10.1167/iovs.12-10222.
Many rhodopsin mutations that cause retinitis pigmentosa produce misfolded rhodopsin proteins that are retained within the endoplasmic reticulum (ER) and cause photoreceptor cell death. Activating transcription factor 6 (ATF6) and protein kinase RNA-like endoplasmic reticulum kinase (PERK) control intracellular signaling pathways that maintain ER homeostasis. The aim of this study was to investigate how ATF6 and PERK signaling affected misfolded rhodopsin in cells, which could identify new molecular therapies to treat retinal diseases associated with ER protein misfolding.
To examine the effect of ATF6 on rhodopsin, wild-type (WT) or mutant rhodopsins were expressed in cells expressing inducible human ATF6f, the transcriptional activator domain of ATF6. Induction of ATF6f synthesis rapidly activated downstream genes. To examine PERK's effect on rhodopsin, WT or mutant rhodopsins were expressed in cells expressing a genetically altered PERK protein, Fv2E-PERK. Addition of the dimerizing molecule (AP20187) rapidly activated Fv2E-PERK and downstream genes. By use of these strategies, it was examined how selective ATF6 or PERK signaling affected the fate of WT and mutant rhodopsins.
ATF6 significantly reduced T17M, P23H, Y178C, C185R, D190G, K296E, and S334ter rhodopsin protein levels in the cells with minimal effects on monomeric WT rhodopsin protein levels. By contrast, the PERK pathway reduced both levels of WT, mutant rhodopsins, and many other proteins in the cell.
This study indicates that selectively activating ATF6 or PERK prevents mutant rhodopsin from accumulating in cells. ATF6 signaling may be especially useful in treating retinal degenerative diseases arising from rhodopsin misfolding by preferentially clearing mutant rhodopsin and abnormal rhodopsin aggregates.
许多导致视网膜色素变性的视紫红质突变会产生错误折叠的视紫红质蛋白，这些蛋白滞留在内质网（ER）中，导致光感受器细胞死亡。激活转录因子 6（ATF6）和蛋白激酶 RNA 样内质网激酶（PERK）控制维持 ER 稳态的细胞内信号通路。本研究旨在探讨 ATF6 和 PERK 信号如何影响细胞内错误折叠的视紫红质，这可能为治疗与 ER 蛋白错误折叠相关的视网膜疾病提供新的分子治疗方法。
为了研究 ATF6 对视紫红质的影响，在表达诱导型人 ATF6f 的细胞中表达野生型（WT）或突变型视紫红质，ATF6f 是 ATF6 的转录激活结构域。ATF6f 合成的诱导迅速激活下游基因。为了研究 PERK 对视紫红质的影响，在表达基因改变的 PERK 蛋白 Fv2E-PERK 的细胞中表达 WT 或突变型视紫红质。二聚化分子（AP20187）的添加迅速激活 Fv2E-PERK 和下游基因。通过这些策略，研究了选择性 ATF6 或 PERK 信号如何影响 WT 和突变型视紫红质的命运。
ATF6 显著降低了 T17M、P23H、Y178C、C185R、D190G、K296E 和 S334ter 视紫红质蛋白水平，对单体 WT 视紫红质蛋白水平的影响最小。相比之下，PERK 通路降低了细胞中 WT、突变型视紫红质和许多其他蛋白的水平。
本研究表明，选择性激活 ATF6 或 PERK 可防止突变型视紫红质在细胞中积累。ATF6 信号可能特别有用，可通过优先清除突变型视紫红质和异常视紫红质聚集体来治疗由视紫红质错误折叠引起的视网膜退行性疾病。
