Dataset Information

Deep Mutational Scan of an SCN5A Voltage Sensor.

ABSTRACT:

Background

Variants in ion channel genes have classically been studied in low throughput by patch clamping. Deep mutational scanning is a complementary approach that can simultaneously assess function of thousands of variants.

Methods

We have developed and validated a method to perform a deep mutational scan of variants in SCN5A, which encodes the major voltage-gated sodium channel in the heart. We created a library of nearly all possible variants in a 36 base region of SCN5A in the S4 voltage sensor of domain IV and stably integrated the library into HEK293T cells.

Results

In preliminary experiments, challenge with 3 drugs (veratridine, brevetoxin, and ouabain) could discriminate wild-type channels from gain- and loss-of-function pathogenic variants. High-throughput sequencing of the pre- and postdrug challenge pools was used to count the prevalence of each variant and identify variants with abnormal function. The deep mutational scan scores identified 40 putative gain-of-function and 33 putative loss-of-function variants. For 8 of 9 variants, patch clamping data were consistent with the scores.

Conclusions

These experiments demonstrate the accuracy of a high-throughput in vitro scan of SCN5A variant function, which can be used to identify deleterious variants in SCN5A and other ion channel genes.

SUBMITTER: Glazer AM

PROVIDER: S-EPMC7031040 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature

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Publications

Deep Mutational Scan of an <i>SCN5A</i> Voltage Sensor.

Glazer Andrew M AM Kroncke Brett M BM Matreyek Kenneth A KA Yang Tao T Wada Yuko Y Shields Tiffany T Salem Joe-Elie JE Fowler Douglas M DM Roden Dan M DM

Circulation. Genomic and precision medicine 20200112 1

<h4>Background</h4>Variants in ion channel genes have classically been studied in low throughput by patch clamping. Deep mutational scanning is a complementary approach that can simultaneously assess function of thousands of variants.<h4>Methods</h4>We have developed and validated a method to perform a deep mutational scan of variants in <i>SCN5A</i>, which encodes the major voltage-gated sodium channel in the heart. We created a library of nearly all possible variants in a 36 base region of <i> ...[more]

PMID: 31928070

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