The State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, #7 Jinsui Road, Guangzhou, Guangdong 510230, China.
Key Laboratory of Protein Chemistry and Developmental Biology, College of Life Sciences, Hunan Normal University, Changsha, Hunan 410081, China.
Curr Mol Med. 2018;18(8):516-522. doi: 10.2174/1566524019666190112144436.
It is well established now that protein sumoylation acts as an important regulatory mechanism mediating control of ocular development through regulation of multiple transcription factors. Yet the functional mechanisms of each factor modulated remain to be further explored using the available in vitro systems. In this regard, various ocular cell lines including HLE, FHL124, αTN4-1, N/N1003A and ARPE-19 have been demonstrated to be useful for biochemical and molecular analyses of normal physiology and pathogenesis. We have recently examined that these cell lines express a full set of sumoylation enzymes E1, E2 and E3. Following this study, here we have examined the localization of these enzymes and determined their differential localization patterns in these major ocular cell lines.
The 5 major ocular cell lines were cultured in Dulbecco's modified Eagle's medium (DMEM) containing fetal bovine serum (FBS) or rabbit serum (RBS) and 1% Penicillin- Streptomycin. The localization of the 3 major sumoylation enzymes in the 5 major ocular cell lines were determined with immunohistochemistry. The images were captured with a Zeiss LSM 880 confocal microscope.
we have obtained the following results: 1) The sumoylation enzymes SAE1, UBC9 and PIAS1 are distributed in both nucleus and cytoplasm, with a much higher level concentrated in the nucleus and the neighboring cellular organelle zone in all cell lines; 2) The sumoylation enzyme UBA2 was highly concentrated in both cytoplasm membrane, cytoskeleton and nucleus of all cell lines; 3) The ligase E3, RanBP2 was exclusively localized in the nucleus with homogeneous distribution.
Our results for the first time established the differential localization patterns of the three types of sumoylation enzymes in 5 major ocular cell lines. Our establishment of the differential localization patterns of the three types of sumoylation enzymes in these cell lines help to predict their functional importance of sumoylation in the vision system. Together, our results demonstrate that these cell lines can be used for assay systems to explore the functional mechanisms of sumoylation mediating ocular development and pathogenesis.
现在已经证实，蛋白质 SUMO 化作为一种重要的调节机制，通过调节多种转录因子来介导对眼部发育的控制。然而，利用现有的体外系统，每个被调节的因子的功能机制仍有待进一步探索。在这方面，各种眼细胞系，包括 HLE、FHL124、αTN4-1、N/N1003A 和 ARPE-19，已被证明可用于正常生理和发病机制的生化和分子分析。我们最近研究发现，这些细胞系表达了一套完整的 SUMO 化酶 E1、E2 和 E3。在这项研究之后，我们在这里检查了这些酶的定位，并确定了它们在这些主要眼细胞系中的差异定位模式。
将 5 种主要的眼细胞系在含有胎牛血清（FBS）或兔血清（RBS）和 1%青霉素-链霉素的 Dulbecco 改良 Eagle 培养基（DMEM）中培养。用免疫组织化学法测定 5 种主要眼细胞系中 3 种主要 SUMO 化酶的定位。图像用蔡司 LSM 880 共聚焦显微镜捕获。
我们得到了以下结果：1）SUMO 化酶 SAE1、UBC9 和 PIAS1 分布在核和细胞质中，在所有细胞系中，核内和邻近的细胞器区集中了更高水平的 SUMO 化酶；2）SUMO 化酶 UBA2 高度集中在细胞质膜、细胞骨架和所有细胞系的核中；3）连接酶 E3、RanBP2 仅定位于核内，分布均匀。
我们的研究首次确定了 5 种主要眼细胞系中 3 种 SUMO 化酶的差异定位模式。我们在这些细胞系中确定了 3 种 SUMO 化酶的差异定位模式，有助于预测 SUMO 化在视觉系统中的功能重要性。总之，我们的研究结果表明，这些细胞系可用于检测系统，以探索 SUMO 化介导的眼部发育和发病机制的功能机制。
