脊椎动物眼晶状体是一种不寻常的器官,其大多数细胞缺乏细胞核和替代衰老蛋白质的能力。小的热休克蛋白α-晶状体蛋白进化成为这种晶状体的关键成分,可能是因为它们能够防止老化蛋白质聚集,否则会导致晶状体不透明。大多数脊椎动物表达两种α-晶状体蛋白,αA-和αB-晶状体蛋白,每个突变都与人类白内障有关。在小鼠敲除模型中,仅αA-晶状体蛋白的缺失导致早期晶状体白内障。我们使用斑马鱼作为模型系统来研究α-晶状体蛋白在晶状体发育过程中的作用。有趣的是,而斑马鱼表达一种晶状体特异性αA-晶状体蛋白基因(cryaa),它们表达两个αB-晶状体蛋白基因,一种进化的晶状体特异性(cryaba),另一种保留其哺乳动物直系同源(cryabb)的广泛表达。在这项研究中,我们使用了所有三个α-晶状体蛋白基因的单个突变斑马鱼系,以确定其丢失对年龄相关性白内障的影响。令人惊讶的是,与老鼠敲除模型不同,我们发现,与24个月大的无cryaa鱼相比,cryaba基因的丢失导致晶状体不透明度增加。αA-晶状体蛋白的缺失并没有增加白内障的患病率。我们还使用单细胞RNA-Seq和RT-qPCR数据显示斑马鱼α-晶状体蛋白在受精后5到10天(dpf)的晶状体表达发生了变化,5和6个dpf镜头几乎只表达cryaa,10dpf后,cryaba和cryabb的表达变得更加突出。这些数据表明,cryaa是早期晶状体发育过程中的主要α-晶状体蛋白,而在镜片老化过程中,对cryaba的保护作用变得更加重要。本研究首次量化野生型斑马鱼的白内障患病率,显示大约25%的鱼在18个月大时出现晶状体混浊。三种α-晶状体蛋白突变体均未显示出其余两种晶状体蛋白表达的代偿性增加,或在丰富的βB1-晶状体蛋白中。总的来说,这些发现表明斑马鱼晶状体发育过程中单个α-晶状体蛋白的功能重要性发生了个体发育变化。我们发现晶状体特异性斑马鱼αBa-晶状体蛋白在预防年龄相关性白内障中起着主导作用,这给我们对脊椎动物晶状体进化的理解增加了新的转折。
The vertebrate eye lens is an unusual organ in that most of its cells lack nuclei and the ability to replace aging protein. The small heat shock protein α-crystallins evolved to become key components of this lens, possibly because of their ability to prevent aggregation of aging protein that would otherwise lead to lens opacity. Most vertebrates express two α-crystallins, αA- and αB-
crystallin, and mutations in each are linked to human cataract. In a mouse knockout model only the loss of αA-
crystallin led to early-stage lens cataract. We have used the zebrafish as a model system to investigate the role of α-crystallins during lens development. Interestingly, while zebrafish express one lens-specific αA-
crystallin gene (cryaa), they express two αB-crystallin genes, with one evolving lens specificity (cryaba) and the other retaining the broad expression of its mammalian ortholog (cryabb). In this study we used individual mutant zebrafish lines for all three α-crystallin genes to determine the impact of their loss on age-related cataract. Surprisingly, unlike mouse knockout models, we found that the loss of the αBa-crystallin gene cryaba led to an increase in lens opacity compared to cryaa null fish at 24 months of age. Loss of αA-
crystallin did not increase the prevalence of cataract. We also used single cell RNA-Seq and RT-qPCR data to show a shift in the lens expression of zebrafish α-crystallins between 5 and 10 days post fertilization (dpf), with 5 and 6 dpf lenses expressing cryaa almost exclusively, and expression of cryaba and cryabb becoming more prominent after 10 dpf. These data show that cryaa is the primary α-crystallin during early lens development, while the protective role for cryaba becomes more important during lens aging. This study is the first to quantify cataract prevalence in wild-type zebrafish, showing that lens opacities develop in approximately 25% of fish by 18 months of age. None of the three α-crystallin mutants showed a compensatory increase in the expression of the remaining two crystallins, or in the abundant βB1-
crystallin. Overall, these findings indicate an ontogenetic shift in the functional importance of individual α-crystallins during zebrafish lens development. Our finding that the lens-specific zebrafish αBa-crystallin plays the leading role in preventing age-related cataract adds a new twist to our understanding of vertebrate lens evolution.