Dazzo Emanuela, Leonardi Emanuela, Belluzzi Elisa, Malacrida Sandro, Vitiello Libero, Greggio Elisa, Tosatto Silvio C E, Nobile Carlo
CNR-Neuroscience Institute, Section of Padua, Padova, Italy.
Department of Woman and Child's Health, University of Padua, Padova, Italy.
PLoS Genet. 2016 Oct 19;12(10):e1006376. doi: 10.1371/journal.pgen.1006376. eCollection 2016 Oct.
Autosomal dominant lateral temporal epilepsy (ADTLE) is a focal epilepsy syndrome caused by mutations in the LGI1 gene, which encodes a secreted protein. Most ADLTE-causing mutations inhibit LGI1 protein secretion, and only a few secretion-positive missense mutations have been reported. Here we describe the effects of four disease-causing nonsynonymous LGI1 mutations, T380A, R407C, S473L, and R474Q, on protein secretion and extracellular interactions. Expression of LGI1 mutant proteins in cultured cells shows that these mutations do not inhibit protein secretion. This finding likely results from the lack of effects of these mutations on LGI1 protein folding, as suggested by 3D protein modelling. In addition, immunofluorescence and co-immunoprecipitation experiments reveal that all four mutations significantly impair interaction of LGI1 with the ADAM22 and ADAM23 receptors on the cell surface. These results support the existence of a second mechanism, alternative to inhibition of protein secretion, by which ADLTE-causing LGI1 mutations exert their loss-of-function effect extracellularly, and suggest that interactions of LGI1 with both ADAM22 and ADAM23 play an important role in the molecular mechanisms leading to ADLTE.
常染色体显性遗传性外侧颞叶癫痫（ADTLE）是一种由LGI1基因突变引起的局灶性癫痫综合征，该基因编码一种分泌蛋白。大多数导致ADLTE的突变会抑制LGI1蛋白的分泌，仅有少数分泌阳性的错义突变被报道。在此，我们描述了四种致病的非同义LGI1突变，即T380A、R407C、S473L和R474Q，对蛋白分泌和细胞外相互作用的影响。在培养细胞中表达LGI1突变蛋白表明，这些突变并不抑制蛋白分泌。正如三维蛋白质建模所显示的，这一发现可能是由于这些突变对LGI1蛋白折叠没有影响。此外，免疫荧光和免疫共沉淀实验表明，所有这四种突变均显著损害了LGI1与细胞表面的ADAM22和ADAM23受体之间的相互作用。这些结果支持了一种不同于抑制蛋白分泌的第二种机制的存在，即导致ADLTE的LGI1突变在细胞外发挥其功能丧失效应的机制，并表明LGI1与ADAM22和ADAM23的相互作用在导致ADLTE的分子机制中发挥重要作用。
