A Role for βA3/A1-Crystallin in Type 2 EMT of RPE Cells Occurring in Dry Age-Related Macular Degeneration.
A Role for βA3/A1-Crystallin in Type 2 EMT of RPE Cells Occurring in Dry Age-Related Macular Degeneration.
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DOI：
10.1167/iovs.18-24132
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发表时间：
2018-03-20
影响因子：
4.4
通讯作者：
Sinha D
中科院分区：
文献类型：
--
作者：
Ghosh S;Shang P;Terasaki H;Stepicheva N;Hose S;Yazdankhah M;Weiss J;Sakamoto T;Bhutto IA;Xia S;Zigler JS Jr;Kannan R;Qian J;Handa JT;Sinha D
RPE细胞在干性年龄相关性黄斑变性（AMD）的发展中起主要作用。我们提出了新的证据表明，由Cryba 1基因编码的βA3/A1-晶状体蛋白（一种已知对RPE中溶酶体清除很重要的蛋白质）也在RPE细胞的上皮向间充质转化（EMT）中发挥作用。使用来自干性AMD球体的RPE、在RPE中全面或特异性缺乏Cryba 1的基因工程小鼠、在Cryba 1中具有功能丧失突变的自发突变大鼠（Nuc 1）和黑色素瘤OCM 3细胞系。蛋白质的空间定位用免疫荧光法证明，基因表达水平用定量PCR（qPCR）测定，蛋白质水平用Western印迹法测定。使用伤口愈合和细胞迁移测定来评价细胞移动。采用免疫共沉淀法鉴定βA3/A1-晶体蛋白的结合伴侣。与未分化细胞相比，极化RPE细胞中βA3/A1-晶状体蛋白上调。小鼠和人RPE细胞中βA3/A1-晶状体蛋白的缺失导致Snail和波形蛋白上调、E-钙粘蛋白下调以及细胞迁移增加。βA3/A1-晶状体蛋白与皮质蛋白结合，βA3/A1-晶状体蛋白的丢失导致P-皮质蛋白Y 421增加。与年龄匹配的对照组相比，AMD样本的RPE具有增加的Snail和波形蛋白，以及减少的E-钙粘蛋白。我们引入了一个新的概念，干性AMD的启动诱导的溶酶体清除缺陷的RPE和随后的尝试，RPE细胞，以避免产生的压力，经历EMT。我们证明，βA3/A1-晶状体蛋白是一个潜在的治疗目标，通过再生的溶酶体功能障碍和潜在的，逆转EMT。
The RPE cells have a major role in the development of dry age-related macular degeneration (AMD). We present novel evidence that βA3/A1-crystallin, encoded by the Cryba1 gene, a protein known to be important for lysosomal clearance in the RPE, also has a role in epithelial-to-mesenchymal transition (EMT) of RPE cells. RPE from dry AMD globes, genetically engineered mice lacking Cryba1 globally or specifically in the RPE, spontaneous mutant rats (Nuc1) with a loss-of-function mutation in Cryba1, and the melanoma OCM3 cell line were used. Spatial localization of proteins was demonstrated with immunofluorescence, gene expression levels were determined by quantitative PCR (qPCR), and protein levels by Western blotting. Cell movement was evaluated using wound healing and cell migration assays. Co-immunoprecipitation was used to identify binding partners of βA3/A1-crystallin. βA3/A1-crystallin is upregulated in polarized RPE cells compared to undifferentiated cells. Loss of βA3/A1-crystallin in murine and human RPE cells resulted in upregulation of Snail and vimentin, downregulation of E-cadherin, and increased cell migration. βA3/A1-crystallin binds to cortactin, and loss of βA3/A1-crystallin resulted in increased P-cortactinY421. The RPE from AMD samples had increased Snail and vimentin, and decreased E-cadherin, compared to age-matched controls. We introduced a novel concept of dry AMD initiation induced by lysosomal clearance defects in the RPE and subsequent attempts by RPE cells to avoid the resulting stress by undergoing EMT. We demonstrate that βA3/A1-crystallin is a potential therapeutic target for AMD through rejuvenation of lysosomal dysfunction and potentially, reversal of EMT.
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DOI：
10.1083/jcb.201509020
发表时间：
2016-02-29
期刊：
The Journal of cell biology
影响因子：
--
作者：
Katsunuma S;Honda H;Shinoda T;Ishimoto Y;Miyata T;Kiyonari H;Abe T;Nibu K;Takai Y;Togashi H
通讯作者：
Togashi H
影响因子：
--
作者：
Myong NH
通讯作者：
Myong NH
DOI：
10.1007/978-1-4939-7553-2_15
发表时间：
2018
期刊：
Methods in molecular biology (Clifton, N.J.)
影响因子：
--
作者：
Khristov V;Wan Q;Sharma R;Lotfi M;Maminishkis A;Bharti K
通讯作者：
Bharti K
影响因子：
12.3
作者：
Radeke MJ;Radeke CM;Shih YH;Hu J;Bok D;Johnson LV;Coffey PJ
通讯作者：
Coffey PJ
影响因子：
9
作者：
Grassi G;Di Caprio G;Santangelo L;Fimia GM;Cozzolino AM;Komatsu M;Ippolito G;Tripodi M;Alonzi T
通讯作者：
Alonzi T