Coexpression of formylglycine-generating enzyme is essential for synthesis and secretion of functional arylsulfatase A in a mouse model of metachromatic leukodystrophy
Coexpression of formylglycine-generating enzyme is essential for synthesis and secretion of functional arylsulfatase A in a mouse model of metachromatic leukodystrophy
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DOI：
10.1089/hum.2005.16.929
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发表时间：
2005-08-01
影响因子：
4.2
通讯作者：
Shimada, T
中科院分区：
文献类型：
--
作者：
Takakusaki, Y;Hisayasu, S;Shimada, T
异色性脑白质营养不良（MLD）是一种溶酶体贮积性疾病，涉及遗传性芳基磺化酶a （ASA）的缺乏。这种疾病的特点是进行性脱髓鞘和在中枢和周围神经系统广泛沉积的硫脂。通过血脑屏障直接注射病毒载体是一种可能的基因治疗方法。然而，为了治疗所有的脑细胞，必须从有限数量的转导细胞中分泌足量的功能性ASA。在本研究中，我们测试了甲酰甘氨酸生成酶（FGE）在功能性ASA过表达中的作用。FGE是一种翻译后修饰酶，对激活多种形式的硫酸盐酶（包括ASA）至关重要。用表达ASA和fge的质粒转染COS-7细胞。通过共表达FGE， ASA活性在细胞裂解液中增加了20倍，在条件培养基中增加了70倍。通过基于流体动力学的方法将表达质粒静脉注射到MLD敲除小鼠体内，导致肝脏和血清中ASA活性的显著协同增加。FGE mRNA的Blot杂交分析表明，内源性FGE在人脑中的表达特别低。我们的研究结果表明，在ASA缺陷交叉校正的基础上，FGE的共表达对于MLD的基因治疗是必不可少的。
Metachromatic leukodystrophy (MLD) is a lysosomal storage disorder involving inherited deficiency of arylsulfatase A (ASA). The disease is characterized by progressive demyelination and widespread deposition of sulfatide in both the central and peripheral nervous systems. Direct injection of viral vector through the blood-brain barrier is a possible gene therapy approach to MILD. However, to treat all brain cells, it is essential to secrete a sufficient amount of functional ASA from limited numbers of transduced cells. In the present study, we tested the utility of formylglycine-generating enzyme (FGE) for overexpression of functional ASA. FGE is a posttranslational modifying enzyme essential for activating multiple forms of sulfatases including ASA. COS-7 cells were transfected with ASA- and FGE-expressing plasmids. ASA activity was increased up to 20-fold in cell lysates and 70-fold in conditioned medium by coexpression of FGE. Intravenous injection of the expression plasmids into MLD knockout mice by a hydrodynamics-based procedure resulted in a significant synergistic increase in ASA activity both in liver and serum. Blot hybridization analysis of FGE mRNA demonstrated that the expression of endogenous FGE was particularly low in human brain. Our results suggest, on the basis of cross-correction of ASA deficiency, that coexpression of FGE is essential for gene therapy of MLD.