LOCALIZATION OF GUANYLATE CYCLASE ACTIVATING PROTEIN 2 IN MAMMALIAN RETINAS

鸟苷酸环化酶激活蛋白 2 在哺乳动物视网膜中的定位

• 批准号: 6116397
• 负责人: KRZYSZTOF PALCZEWSKI
• 金额: $ 8.67万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-01 至 2000-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6116397
• 关键词: Primates; animal tissue; bioimaging /biomedical imaging; eye; human tissue; microscopy; nervous system; proteins

When the eyes of vertebrate animals have been exposed to light and then the light is extinguished, rod photoreceptors in the neural retina must re-establish a dark-adapted state. A critical step in this process is the intracellular stimulation of the enzyme guanylate cyclase (GC). GC-activating proteins (GCAP1 and GCAP2) modulate GC activation. GCAP activation of GC is regulated by the concentration of intracellular calcium ions [Ca2+]i. Previous studies have localized GCAP1 to rod and cone outer segments, somata and synaptic terminals. In the current study, we used in situ hybridization and immunocytochemistry to localize GCAP2 in human, monkey, and bovine retinas. In human and monkey retinas, the most intense immunolabeling with anti-GCAP2 antibodies was in the cone inner segments, somata and synaptic terminals and, to a lesser degree, in the rod inner segments and inner retinal neurons. In bovine retinas, the most intense immunolabeling was in the rod inner segments, with weaker labeling of cone myoids, somata and synapses. Additional enzymatic assays using a GCAP2-specific antibody confirmed that GCAP1, but not GCAP2, is the major component that stimulates GC in bovine rod outer segments. These results suggest that although GCAP1 is involved in the Ca2+-sensitive regulation of GC in rod and cone outer segments, GCAP2 may have non-phototransduction functions in photoreceptors and inner retinal neurons. Although the role of GCAP1 and GCAP2 mutations in human retinal diseases has yet to be established, mutations in GC are known to cause a severe retinopathy known as Leber's congenital amaurosis. Individuals with loss of function mutations in GCAPs might be expected to exhibit a similar phenotype.

当脊椎动物的眼睛暴露在光线下，然后光线熄灭时，神经视网膜中的杆状光感受器必须重新建立一种适应黑暗的状态。这个过程的关键步骤是胞内刺激鸟苷酸环化酶（GC）。气相色谱激活蛋白（GCAP1和GCAP2）调节气相色谱激活。GCAP对GC的激活受细胞内钙离子[Ca2+]浓度的调控。以往的研究将GCAP1定位于杆状体和锥体外节、体细胞和突触末端。在本研究中，我们使用原位杂交和免疫细胞化学方法在人、猴和牛视网膜中定位GCAP2。在人和猴视网膜中，抗gcap2抗体的免疫标记最强烈的是在视锥内段、体细胞和突触末端，其次是在视杆内段和视网膜内神经元。在牛视网膜中，视杆内段的免疫标记最强烈，锥体肌样体、体细胞和突触的免疫标记较弱。使用GCAP2特异性抗体的酶促分析证实，GCAP1是刺激牛棒外段GC的主要成分，而不是GCAP2。这些结果表明，尽管GCAP1参与了杆状细胞和锥体细胞外段Ca2+敏感的GC调控，但GCAP2可能在光感受器和视网膜内神经元中具有非光导功能。虽然GCAP1和GCAP2突变在人类视网膜疾病中的作用尚未确定，但已知GC突变可引起称为Leber先天性黑朦的严重视网膜病变。gcap中功能突变丧失的个体可能会表现出类似的表型。