Hadzijahic Nedym, Kim Colin K, Djulbegovic Mak B, Antonietti Michael, Taylor Gonzalez David J, Uversky Vladimir N, Pulido Jose S, Karp Carol L
Bascom Palmer Eye Institute, University of Miami, 900 NW 17th Street, Miami, FL 33136, USA.
Wills Eye Hospital, Thomas Jefferson University, Philadelphia, PA, USA.
J Proteins Proteom. 2025 Jun 19. doi: 10.1007/s42485-025-00188-6.
The human retina is integral to vision, converting light into neural signals through a complex interplay of specialized neuronal cell types. Recent proteomic studies have revealed significant insights into retinal function, yet much of the retina's proteome remains unexplored. Our research focuses on quantifying and characterizing intrinsically disordered proteins (IDPs) and regions (IDRs) within the retina and other ocular structures. These proteins are critical for cellular processes due to their flexible, structure-less nature, allowing for versatile interactions in signaling and regulatory networks. Furthermore, we investigate the phenomenon of liquid-liquid-phase separation (LLPS), a process vital for cellular organization and implicated in various diseases, within the retina proteome.
In this study, we employed a suite of bioinformatics and deep learning tools to analyze protein intrinsic disorder and the propensity for LLPS in proteomes from both healthy and diseased retinas. We utilized the Human Protein Atlas (HPA) as a baseline control, comparing it against the RetNet protein set and samples afflicted by age-related macular degeneration (AMD), glaucoma, and diabetic retinopathy (DR) with and without gliosis. Protein sequences were sourced from the universal protein resource (UniProt) and analyzed for intrinsic disorder using the rapid intrinsic disorder analysis online (RIDAO) platform. Disorder levels and phase separation tendencies were further examined through statistical analyses, including ANOVA and chi-squared tests, to evaluate differences across proteomes. In addition, we assessed the likelihood of proteins to undergo LLPS using predictive tools, such as PSPredictor and ParSe V2, integrating these findings with intrinsic disorder data to draw comprehensive conclusions about the structural dynamics within these proteomes.
The HPA control proteome displayed the highest levels of intrinsic disorder, significantly greater than those observed in disease-specific proteomes, including those affected by AMD, glaucoma, and diabetic retinopathy with and without gliosis. CH-CDF plot analysis revealed distinct structural profiles, with a higher proportion of structured proteins in the HPA and molten globular states prevalent in disease states. Our findings highlight a marked disparity in LLPS propensity, with the HPA proteome and the RetNet Protein Set demonstrating the greatest potential, suggesting a disease-specific alteration in protein interaction dynamics and structural organization.
This study revealed significant variations in protein intrinsic disorder and liquid-LLPS across healthy and diseased retinal proteomes. The highest levels of disorder in the HPA proteome suggest a proteomic flexibility that is critical for normal retinal function. In contrast, the AMD and glaucoma proteomes, with their lower disorder and LLPS propensity, may lack this adaptability, potentially contributing to disease progression. These insights underscore the importance of protein dynamics in retinal disorders and point towards targeted therapies that could manipulate these properties to improve or maintain retinal health.
人类视网膜对于视觉至关重要，它通过特殊神经元细胞类型的复杂相互作用将光转化为神经信号。最近的蛋白质组学研究已揭示了对视网膜功能的重要见解，但视网膜的大部分蛋白质组仍未被探索。我们的研究重点是对视网膜及其他眼部结构中的内在无序蛋白质（IDP）和区域（IDR）进行定量和表征。由于这些蛋白质具有灵活、无结构的性质，它们对细胞过程至关重要，能够在信号传导和调节网络中进行多种相互作用。此外，我们还研究了液-液相分离（LLPS）现象，这一过程对细胞组织至关重要且与多种疾病相关，我们在视网膜蛋白质组中对其进行研究。
在本研究中，我们使用了一系列生物信息学和深度学习工具来分析健康和患病视网膜蛋白质组中蛋白质的内在无序性以及LLPS倾向。我们将人类蛋白质图谱（HPA）用作基线对照，将其与RetNet蛋白质集以及患有年龄相关性黄斑变性（AMD）、青光眼和糖尿病性视网膜病变（DR）（有无神经胶质增生）的样本进行比较。蛋白质序列来自通用蛋白质资源（UniProt），并使用在线快速内在无序分析（RIDAO）平台分析其内在无序性。通过方差分析和卡方检验等统计分析进一步检查无序水平和相分离倾向，以评估不同蛋白质组之间的差异。此外，我们使用PSPredictor和ParSe V2等预测工具评估蛋白质发生LLPS的可能性，并将这些结果与内在无序数据相结合，以得出关于这些蛋白质组内结构动力学的全面结论。
HPA对照蛋白质组显示出最高水平的内在无序性，显著高于在疾病特异性蛋白质组中观察到的水平，包括那些受AMD、青光眼以及有无神经胶质增生的糖尿病性视网膜病变影响的蛋白质组。CH-CDF图分析揭示了不同的结构特征，HPA中结构化蛋白质比例更高，而疾病状态下则普遍存在熔球态。我们的研究结果突出了LLPS倾向的显著差异，HPA蛋白质组和RetNet蛋白质集显示出最大的潜力，这表明蛋白质相互作用动力学和结构组织存在疾病特异性改变。
本研究揭示了健康和患病视网膜蛋白质组中蛋白质内在无序性和液-LLPS的显著差异。HPA蛋白质组中最高水平的无序性表明蛋白质组的灵活性对正常视网膜功能至关重要。相比之下，AMD和青光眼蛋白质组的无序性和LLPS倾向较低，可能缺乏这种适应性，这可能导致疾病进展。这些见解强调了蛋白质动力学在视网膜疾病中的重要性，并指出了可以操纵这些特性以改善或维持视网膜健康的靶向治疗方法。
