Regulation of retinal homeostasis and disease by Fic-mediated AMPylation

项目总结。 蛋白质的正确合成、折叠、修饰和降解对细胞的健康和功能至关重要。 这些过程，统称为蛋白质稳态/蛋白质稳态，随着时间的推移已经演变成 错综复杂的机制，在细胞内进行仔细的调节。未折叠蛋白反应(UPR)是一种细胞 当错误折叠的蛋白质在内质网(ER)中积累时被激活的应激反应。 UPR的激活对细胞的正常功能和健康至关重要；然而，慢性或长期的UPR 导致炎症加剧和细胞凋亡的激活。如果这发生在有丝分裂后细胞中，组织 无法重新生成。因此，当这种情况发生在视网膜的光感受器神经元中时，它会导致不可逆的 失明。慢性或调节失调的UPR与多种视网膜退行性疾病有关；例如 糖尿病视网膜病变，青光眼，Leber先天性黑色素和视网膜色素变性(RP)。对…的调查 导致光感受器退化的UPR的作用可以提供对靶点的重要洞察 寻找治疗视网膜退行性疾病患者的新的治疗途径。众所周知，普遍定期审议 受内质网伴侣蛋白Bip的调节，它既是清除错误折叠蛋白质的分子伴侣 并作为普遍定期审议不同分支机构的监管者。我们第一次发现，这种酶 可以通过Bip的翻译后修饰(AMP/去AMP)来调节UPR。这 提示Fic介导的BiP的AMP化作用是UPR的分子变阻器。为了支持这一点，我们 研究发现，果蝇体内FIC的丢失会导致视力障碍，并改变视网膜和大脑中UPR的活性 由于暴露在连续的光线下而触发的眼睛的板层。我们已经生成了一种新的老鼠模型 我们可以研究Fic介导的Bip AMP化在哺乳动物视网膜中的确切作用。我们 假设通过BiP的Fic AMP化来调节UPR对于防止光感受器死亡是必要的 和视力丧失。我们将解决以下问题：1)Fic-/-小鼠是否表现出UPR激活的改变 正常生理条件下的视网膜，以及2)Fic-/-小鼠容易受到UPR相关的损伤 在压力和疾病状态下？该项目的调查结果将开发有价值的工具，用于监测和 在正常生理状态下哺乳动物视网膜细胞中没有Fic的情况下确定UPR 衰老和视网膜退行性疾病状态。发现Fic在内质网调控中的作用 哺乳动物视网膜的动态平衡可以为未来潜在的治疗方法提供细胞靶点。 治疗或预防内质网应激相关的光感受器细胞死亡和视力丧失。

PROJECT SUMMARY. The proper synthesis, folding, modification and degradation of proteins is vital to cellular health and function. These processes, known collectively as protein homeostasis/proteostasis, have evolved over time to have intricate mechanisms in place for careful regulation in the cell. The unfolded protein response (UPR) is a cellular stress response that is activated when misfolded proteins accumulate in the endoplasmic reticulum (ER). Activation of the UPR is critical for normal cellular function and health; however, a chronic or prolonged UPR results in elevated inflammation and the activation of apoptosis. If this occurs in post-mitotic cells, the tissue cannot be regenerated. Thus, when this occurs in the photoreceptor neurons of the retina, it causes irreversible blindness. Chronic or dysregulated UPR has been linked to a variety of retinal degenerative diseases; such as diabetic retinopathy, glaucoma, Leber congenital amaurosis, and retinitis pigmentosa (RP). Investigation into the role of the UPR that leads to photoreceptor degeneration can provide important insight into targets for novel therapeutic avenues to treat patients with retinal degenerative diseases. The UPR is known to be regulated by the ER chaperone BiP, which acts as both a molecular chaperone to clear misfolded proteins and as a regulator of the different branches of the UPR. We discovered, for first time, that the enzyme Fic can modulate the UPR via post-translational modification (AMPylation/deAMPylation) of BiP. This indicates that Fic-mediated AMPylation of BiP acts as a molecular rheostat for the UPR. In support of this, we found that a loss of fic in Drosophila leads to vision defects and altered UPR activation in the both the retina and lamina of the eye triggered by exposure to continuous light. We have generated a novel mouse model in which we can study the precise role of Fic-mediated BiP AMPylation in the mammalian retina. We hypothesize that the regulation of the UPR via Fic AMPylation of BiP is necessary to prevent photoreceptor death and vision loss. We will address the following questions: 1) do Fic-/- mice exhibit altered UPR activation in the retina under normal physiological conditions, and 2) are Fic-/- mice predisposed to UPR-associated damage under stress and disease states? The findings of this project will develop valuable tools for monitoring and defining the UPR in the absence of Fic in mammalian retinal cells, both during normal physiological aging and in retinal degenerative disease states. Discovering the role Fic plays in the regulation of ER homeostasis in the mammalian retina can provide insight into cellular targets for potential future therapeutics to treat or prevent ER stress-related photoreceptor cell death and vision loss.