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The connexin 46 mutant (V44M) impairs gap junction function causing congenital cataract

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Abstract

Connexin 46 (Cx46) is important for gap junction channels formation which plays crucial role in the preservation of lens homeostasis and transparency. Previously, we have identified a missense mutation (p.V44M) of Cx46 in a congenital cataract family. This study aims at dissecting the potential pathogenesis of the causative mutant of cataract. Plasmids carrying wild-type (wt) and mutant (V44M) of Cx46 were constructed and expressed in Hela cells respectively. Western blotting and fluorescence microscopy were applied to analyse the expression and subcellular localization of recombinant proteins, respectively. Scrape loading dye transfer experiment was performed to detect the transfer capability of gap junction channels among cells expressed V44M mutant. The results demonstrated that in transfected Hela cells, both wt-Cx46 and Cx46 V44M were localized abundantly in the plasma membrane. No significant difference was found between the protein expressions of the two types of Cx46. The fluorescent localization assay revealed the plaque formation, significantly reduced in the cells expressing Cx46 V44M. Immunoblotting analysis demonstrated that formation of Triton X-100 insoluble complex decreased obviously in mutant Cx46. Additionally, the scrape-loading dye-transfer experiment showed a lower dye diffusion distance of Cx46 V44M cells, which indicates that the gap junction intercellular communication activity was aberrant. Human Cx46 V44M mutant causing cataracts result in abnormally decreased formation of gap junction plaques and impaired gap junction channel function.

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Acknowledgements

This work was supported by grants from the National Natural Science Foundation of China (grant no. 31601116); the China Postdoctoral Science Foundation (grant no. 2016M601447) and the Natural Science Foundation of Heilongjiang Province of China (grant no. QC2014C120).

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    Authors and Affiliations

    1. Department of Ophthalmology, Hongqi Hospital of Mudanjiang Medical College, Mudanjiang, 157000, Heilongjiang Province, People’s Republic of China

      Lijuan Chen, Sijia Li & Shanshan Hu

    2. Department of Genetics, National Research Institute for Family Planning, Beijing, 100081, People’s Republic of China

      Dongmei Su, Lina Guan, Cuige Shi & Xu Ma

    3. Department of Immunology, Basic Medical College of Harbin Medical University, Harbin, 150081, People’s Republic of China

      Dianjun Li & Shanshan Hu

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    1. Lijuan Chen
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    Correspondence to Shanshan Hu or Xu Ma.

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    Corresponding editor: Kunal Ray

    Lijuan Chen, Dongmei Su, Shanshan Hu and Xu Ma contributed equally to this work.

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    Chen, L., Su, D., Li, S. et al. The connexin 46 mutant (V44M) impairs gap junction function causing congenital cataract. J Genet 96, 969–976 (2017). https://doi.org/10.1007/s12041-017-0861-0

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