杆和锥磷酸二酯酶6(PDE6)是脊椎动物光转导途径的关键效应酶。rodPDE6由两个催化亚基PDE6α和PDE6β以及两个相同的抑制性PDE6γ亚基组成,而视锥PDE6由两个相同的PDE6α催化亚基和两个相同的视锥特异性PDE6γ抑制亚基组成。尽管它们在调节cGMP水平方面具有突出的功能,因此具有杆状和锥形光响应特性,它是不知道如何每个亚基有助于杆和锥体之间的功能差异。在这项研究中,我们产生了一个rd10/cpfl1小鼠模型,缺乏杆PDE6β和锥PDE6α\'亚基。视杆细胞和视锥细胞都随着年龄的增长而退化,所有PDE6亚基在rd10/cpfl1小鼠中都会降解。我们通过腺相关病毒(AAV)介导的传递在rd10/cpfl1小鼠的视杆和视锥中表达视锥PDE6α\',组成型小鸡β-肌动蛋白启动子。我们表明PDE6α的表达可以挽救rd10/cpfl1小鼠的杆功能,通过恢复内源性棒PDE6γ和形成功能性PDE6α\'γ复合物来恢复棒光敏感性。然而,仅在补充视锥PDE6α和PDE6γ亚基后,才能获得改善的适光视锥反应,而不是单独通过PDE6α处理。我们观察到注射PDE6α+PDE6γ的眼睛中PDE6α水平相对于单独接受PDE6α的眼睛增加了两倍。尽管同时存在PDE6γ和PDE6γ,大多数PDE6α与PDE6γ形成了功能复合物,表明PDE6α对PDE6γ的缔合亲和力高于对PDE6γ的缔合亲和力。这些结果表明PDE6γ的存在增强了圆锥PDE6的组装并增强了其稳定性。我们的发现对PDE6α相关色盲的基因治疗具有重要意义。
Rod and cone phosphodiesterase 6 (PDE6) are key effector enzymes of the vertebrate phototransduction pathway. Rod PDE6 consists of two catalytic subunits PDE6α and PDE6β and two identical inhibitory PDE6γ subunits, while cone PDE6 is composed of two identical PDE6α\' catalytic subunits and two identical cone-specific PDE6γ\' inhibitory subunits. Despite their prominent function in regulating cGMP levels and therefore rod and cone light response properties, it is not known how each subunit contributes to the functional differences between rods and cones. In this study, we generated an rd10/
cpfl1 mouse model lacking rod PDE6β and cone PDE6α\' subunits. Both rod and cone photoreceptor cells are degenerated with age and all PDE6 subunits degrade in rd10/
cpfl1 mice. We expressed cone PDE6α\' in both rods and cones of rd10/
cpfl1 mice by adeno-associated virus (AAV)-mediated delivery driven by the ubiquitous, constitutive small chicken β-actin promoter. We show that expression of PDE6α\' rescues rod function in rd10/
cpfl1 mice, and the restoration of rod light sensitivity is attained through restoration of endogenous rod PDE6γ and formation of a functional PDE6α\'γ complex. However, improved photopic cone responses were achieved only after supplementation of both cone PDE6α\' and PDE6γ\' subunits but not by PDE6α\' treatment alone. We observed a two fold increase of PDE6α\' levels in the eyes injected with both PDE6α\' plus PDE6γ\' relative to eyes receiving PDE6α\' alone. Despite the presence of both PDE6γ\' and PDE6γ, the majority of PDE6α\' formed functional complexes with PDE6γ\', suggesting that PDE6α\' has a higher association affinity for PDE6γ\' than for PDE6γ. These results suggest that the presence of PDE6γ\' augments cone PDE6 assembly and enhances its stability. Our finding has important implication for gene therapy of PDE6α\'-associated achromatopsia.