Lens Epithelial Explants Treated with Vitreous Humor Undergo Alterations in Chromatin Landscape with Concurrent Activation of Genes Associated with Fiber Cell Differentiation and Innate Immune Response.
Lens Epithelial Explants Treated with Vitreous Humor Undergo Alterations in Chromatin Landscape with Concurrent Activation of Genes Associated with Fiber Cell Differentiation and Innate Immune Response.
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晶状体上皮外植体由在其天然基底膜（晶状体囊）上体外培养的晶状体上皮细胞组成。生物学家使用晶状体上皮外植体来研究许多不同的细胞过程，包括晶状体纤维细胞分化。在这些研究中，纤维分化通常通过细胞伸长和晶状体纤维细胞原位表达的一些蛋白质的表达来测量。从晶状体上皮外植体中收集染色质和 RNA，该晶状体上皮外植体在未补充的培养基或含有 50% 牛玻璃体液的培养基中培养一或五天。从外植体中制备用于 ATAC 测序的染色质和用于 RNA 测序的 RNA，以分别评估可接近染色质的区域并定量测量基因表达。玻璃体液增加了与纤维分化相关的基因启动子区域的染色质可及性，令人惊讶的是，与免疫反应相关，这与这些基因的转录水平增加有关。相比之下，玻璃体对晶状体上皮中高表达基因的可及性几乎没有影响，尽管其 mRNA 转录物显着减少。对差异可及区域的公正分析揭示了 RUNX、SOX 和 TEAD 转录因子的顺式调控基序的富集，这些转录因子可能驱动响应玻璃体的差异基因表达。
Lens epithelial explants are comprised of lens epithelial cells cultured in vitro on their native basement membrane, the lens capsule. Biologists have used lens epithelial explants to study many different cellular processes including lens fiber cell differentiation. In these studies, fiber differentiation is typically measured by cellular elongation and the expression of a few proteins characteristically expressed by lens fiber cells in situ. Chromatin and RNA was collected from lens epithelial explants cultured in either un-supplemented media or media containing 50% bovine vitreous humor for one or five days. Chromatin for ATAC-sequencing and RNA for RNA-sequencing was prepared from explants to assess regions of accessible chromatin and to quantitatively measure gene expression, respectively. Vitreous humor increased chromatin accessibility in promoter regions of genes associated with fiber differentiation and, surprisingly, an immune response, and this was associated with increased transcript levels for these genes. In contrast, vitreous had little effect on the accessibility of the genes highly expressed in the lens epithelium despite dramatic reductions in their mRNA transcripts. An unbiased analysis of differentially accessible regions revealed an enrichment of cis-regulatory motifs for RUNX, SOX and TEAD transcription factors that may drive differential gene expression in response to vitreous.
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