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Updated: May 6, 2026
Targeted Next-generation Sequencing and Bioinformatics Pipeline to Evaluate Genetic Determinants of Constitutional Disease
Published on: April 4, 2018
Functional analysis of NPHS1 mutations in Japanese patients
Takayuki Miyai1, Kunihiko Aya2, Masanori Takaiwa1
1Department of Pediatrics, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Okayama, Japan.
Background:
Many mutations in the NPHS1 gene were detected among patients with congenital nephrotic syndrome. Functional analysis of those mutations was done with a stable-expression cell line. Nevertheless, establishing such a cell line is time-consuming.
Methods And Results:
We established an easier method using automatic counting software for functional analysis with transient-transfection cells rather than a stable-expression cell line. We demonstrated maltrafficking to the plasma membrane of abnormal nephrin for immunostaining on transient-expression cells by comparison without Triton X (detecting proteins on the cell membrane only) and with Triton X (detecting proteins both on the cell membrane and inside the cell cytoplasm). We obtained relevant results with data obtained previously using a stable-expression cell line. Furthermore, we conducted functional analysis of NPHS1 mutations in Japanese patients with congenital nephrotic syndrome using this simple method, which revealed that all pathogenic mutations impaired trafficking to the protein plasma membrane.
Conclusions:
Functional analysis using transient-expression cells with automatic counting software was useful to demonstrate maltrafficking to the plasma membrane of a protein. All pathogenic mutations detected in Japanese patients impaired trafficking to the protein plasma membrane.
