Sacristan-Reviriego Almudena, Bellingham James, Prodromou Chrisostomos, Boehm Annika N, Aichem Annette, Kumaran Neruban, Bainbridge James, Michaelides Michel, van der Spuy Jacqueline
UCL Institute of Ophthalmology, London EC1V 9EL, UK.
Genome Damage and Stability Centre, University of Sussex, Brighton, East Sussex BN1 9RQ, UK.
Hum Mol Genet. 2017 Nov 15;26(22):4465-4480. doi: 10.1093/hmg/ddx334.
Biallelic mutations in the photoreceptor-expressed aryl hydrocarbon receptor interacting protein-like 1 (AIPL1) are associated with autosomal recessive Leber congenital amaurosis (LCA), the most severe form of inherited retinopathy in early childhood. AIPL1 functions as a photoreceptor-specific co-chaperone that interacts with the molecular chaperone HSP90 to facilitate the stable assembly of the retinal cyclic GMP (cGMP) phosphodiesterase (PDE6) holoenzyme. In this study, we characterized the functional deficits of AIPL1 variations, some of which induce aberrant pre-mRNA AIPL1 splicing leading to the production of alternative AIPL1 isoforms. We investigated the ability of the AIPL1 variants to mediate an interaction with HSP90 and modulate the rod cGMP PDE6 stability and activity. Our data revealed that both the FK506 binding protein (FKBP)-like domain and the tetratricopeptide repeat (TPR) domain of AIPL1 are required for interaction with HSP90. We further demonstrate that AIPL1 significantly modulates the catalytic activity of heterologously expressed rod PDE6. Although the N-terminal FKBP-like domain of AIPL1 binds the farnesylated PDE6α subunit through direct interaction with the farnesyl moiety, mutations compromising the integrity of the C-terminal TPR domain of AIPL1 also failed to modulate PDE6 activity efficiently. These AIPL1 variants moreover failed to promote the HSP90-dependent stabilization of the PDE6α subunit in the cytosol. In summary, we have successfully validated the disease-causing status of the AIPL1 variations in vitro. Our findings provide insight into the mechanism underlying the co-chaperone role of AIPL1 and will be critical for ensuring an early and effective diagnosis of AIPL1 LCA patients.
在光感受器中表达的芳烃受体相互作用蛋白样1（AIPL1）的双等位基因突变与常染色体隐性遗传性莱伯先天性黑蒙（LCA）相关，这是幼儿期最严重的遗传性视网膜病变形式。AIPL1作为一种光感受器特异性共伴侣蛋白，与分子伴侣HSP90相互作用，以促进视网膜环磷酸鸟苷（cGMP）磷酸二酯酶（PDE6）全酶的稳定组装。在本研究中，我们对AIPL1变异的功能缺陷进行了表征，其中一些变异会诱导异常的前体mRNA AIPL1剪接，导致产生替代性AIPL1异构体。我们研究了AIPL1变体介导与HSP90相互作用并调节视杆细胞cGMP PDE6稳定性和活性的能力。我们的数据显示，AIPL1的FK506结合蛋白（FKBP）样结构域和四肽重复（TPR）结构域都是与HSP90相互作用所必需的。我们进一步证明，AIPL1显著调节异源表达的视杆细胞PDE6的催化活性。虽然AIPL1的N端FKBP样结构域通过与法尼基部分直接相互作用结合法尼基化的PDE6α亚基，但损害AIPL1 C端TPR结构域完整性的突变也无法有效调节PDE6活性。此外，这些AIPL1变体未能促进PDE6α亚基在细胞质中依赖HSP90的稳定。总之，我们在体外成功验证了AIPL1变异的致病状态。我们的发现为AIPL1共伴侣作用的潜在机制提供了见解，对于确保早期有效诊断AIPL1 LCA患者至关重要。
