Pharmacological rescue of carnitine transport in primary carnitine deficiency
Pharmacological rescue of carnitine transport in primary carnitine deficiency
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DOI：
10.1002/humu.20314
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发表时间：
2006-06-01
期刊：
影响因子：
3.9
通讯作者：
Longo, Nicola
中科院分区：
文献类型：
--
作者：
Filippo, Cristina Amat di San;Pasquali, Marzia;Longo, Nicola
原发性肉毒碱缺乏症是由编码OCTN 2肉毒碱转运蛋白的SLC 22 A3基因的异质性突变引起的隐性疾病。在这里，我们扩展到八个新的家庭这种疾病的突变分析。为了确定错义突变损害肉毒碱转运的机制，用绿色荧光蛋白标记OCTN 2转运蛋白并在CHO细胞中表达。共聚焦显微镜分析表明，几个错义突变体（M1 I，R169 W，T232 M，G242 V，S280 F，R282 Q，W283 R，A301 D，W35111，R399 Q，T440 M，E452 K和T468 R）正常成熟到质膜。相比之下，其他突变（包括R19 P、Delta F22、R83 L、S280 F、P398 L、Y 447 C和A142 S/R488 H）导致突变型0 CTN 2转运蛋白在细胞质中的显著保留。成熟至质膜失败是影响膜转运蛋白/离子通道的疾病（包括囊性纤维化）的常见机制。为了纠正这一缺陷，我们测试了降低内质网（ER）中蛋白质降解效率的药物（苯丁酸盐，姜黄素）或能够结合OCTN 2肉毒碱转运蛋白的药物（维拉帕米，奎尼丁）是否可以改善肉毒碱转运。长时间孵育苯丁酸，奎尼丁，维拉帕米部分刺激肉毒碱运输，而姜黄素是无效的。这些结果表明，OCTN 2突变可以通过损害成熟的转运蛋白质膜影响肉毒碱运输。药物治疗可以有效地部分恢复突变转运蛋白的活性。
Primary carnitine deficiency is a recessive disorder caused by heterogeneous mutations in the SLC22A3 gene encoding the OCTN2 carnitine transporter. Here we extend mutational analysis to eight new families with this disorder. To determine the mechanism by which missense mutations impaired carnitine transport, the OCTN2 transporter was tagged with the green fluorescent protein and expressed in CHO cells. Analysis by confocal microscopy indicated that several missense mutants (M1I, R169W, T232 M, G242 V, S280F, R282Q, W283R, A301D, W35111, R399Q, T440M, E452K, and T468R) matured normally to the plasma membrane. By contrast, other mutations (including R19P, Delta F22, R83L, S280F, P398L, Y447C, and A142S/R488H) caused significant retention of the mutant OCTN2 transporter in the cytoplasm. Failed maturation to the plasma membrane is a common mechanism in disorders affecting membrane transporters/ion channels, including cystic fibrosis. To correct this defect, we tested whether drugs reducing the efficiency of protein degradation in the endoplasmic reticulum (ER) (phenylbutyrate, curcumin) or capable of binding the OCTN2 carnitine transporter (verapamil, quinidine) could improve carnitine transport. Prolonged incubation with phenylbutyrate, quinidine, and verapamil partially stimulated carnitine transport, while curcumin was ineffective. These results indicate that OCTN2 mutations can affect carnitine transport by impairing maturation of transporters to the plasma membrane. Pharmacological therapy can be effective in partially restoring activity of mutant transporters.