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Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines

Asparagine synthetase (ASNS) catalyzes the synthesis of asparagine (Asn) from aspartate and glutamine. Biallelic mutations in the ASNS gene result in ASNS Deficiency (ASNSD). Children with ASNSD exhibit congenital microcephaly, epileptic‐like seizures, and continued brain atrophy, often leading to p...

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Autores principales: Staklinski, Stephen J., Chang, Mario C., Ahrens‐Nicklas, Rebecca C., Kaur, Shagun, Stefanatos, Arianna K., Dudenhausen, Elizabeth E., Merritt, Matthew E., Kilberg, Michael S.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley & Sons, Inc. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9981421/
https://www.ncbi.nlm.nih.gov/pubmed/36873094
http://dx.doi.org/10.1002/jmd2.12356
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author Staklinski, Stephen J.
Chang, Mario C.
Ahrens‐Nicklas, Rebecca C.
Kaur, Shagun
Stefanatos, Arianna K.
Dudenhausen, Elizabeth E.
Merritt, Matthew E.
Kilberg, Michael S.
author_facet Staklinski, Stephen J.
Chang, Mario C.
Ahrens‐Nicklas, Rebecca C.
Kaur, Shagun
Stefanatos, Arianna K.
Dudenhausen, Elizabeth E.
Merritt, Matthew E.
Kilberg, Michael S.
author_sort Staklinski, Stephen J.
collection PubMed
description Asparagine synthetase (ASNS) catalyzes the synthesis of asparagine (Asn) from aspartate and glutamine. Biallelic mutations in the ASNS gene result in ASNS Deficiency (ASNSD). Children with ASNSD exhibit congenital microcephaly, epileptic‐like seizures, and continued brain atrophy, often leading to premature mortality. This report describes a 4‐year‐old male with global developmental delay and seizures with two novel mutations in the ASNS gene, c.614A > C (maternal) and c.1192dupT (paternal) encoding p.H205P and p.Y398Lfs*4 variants, respectively. We employed the novel use of immortalized lymphoblastoid cell lines (LCL) to show that the proliferation of the heterozygotic parental LCL was not severely affected by culture in Asn‐free medium, but growth of the child's cells was suppressed by about 50%. Asn production by the LCL from both the father and the child was significantly decreased relative to the mother's cells. mRNA and protein analysis of the paternal LCL cells for the Y398Lfs*4 variant revealed reductions in both. Attempts to ectopically express the truncated Y398Lfs*4 variant in either HEK293T or ASNS‐null cells resulted in little or no detectable protein. Expression and purification of the H205P variant from HEK293T cells revealed enzymatic activity similar to wild‐type ASNS. Stable expression of WT ASNS rescued the growth of ASNS‐null JRS cells in Asn‐free medium and the H205P variant was only slightly less effective. However, the Y398Lfs*4 variant appeared to be unstable in JRS cells. These results indicate that co‐expression of the H205P and Y398Lfs*4 variants leads to a significant reduction in Asn synthesis and cellular growth.
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spelling pubmed-99814212023-03-04 Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines Staklinski, Stephen J. Chang, Mario C. Ahrens‐Nicklas, Rebecca C. Kaur, Shagun Stefanatos, Arianna K. Dudenhausen, Elizabeth E. Merritt, Matthew E. Kilberg, Michael S. JIMD Rep Research Reports Asparagine synthetase (ASNS) catalyzes the synthesis of asparagine (Asn) from aspartate and glutamine. Biallelic mutations in the ASNS gene result in ASNS Deficiency (ASNSD). Children with ASNSD exhibit congenital microcephaly, epileptic‐like seizures, and continued brain atrophy, often leading to premature mortality. This report describes a 4‐year‐old male with global developmental delay and seizures with two novel mutations in the ASNS gene, c.614A > C (maternal) and c.1192dupT (paternal) encoding p.H205P and p.Y398Lfs*4 variants, respectively. We employed the novel use of immortalized lymphoblastoid cell lines (LCL) to show that the proliferation of the heterozygotic parental LCL was not severely affected by culture in Asn‐free medium, but growth of the child's cells was suppressed by about 50%. Asn production by the LCL from both the father and the child was significantly decreased relative to the mother's cells. mRNA and protein analysis of the paternal LCL cells for the Y398Lfs*4 variant revealed reductions in both. Attempts to ectopically express the truncated Y398Lfs*4 variant in either HEK293T or ASNS‐null cells resulted in little or no detectable protein. Expression and purification of the H205P variant from HEK293T cells revealed enzymatic activity similar to wild‐type ASNS. Stable expression of WT ASNS rescued the growth of ASNS‐null JRS cells in Asn‐free medium and the H205P variant was only slightly less effective. However, the Y398Lfs*4 variant appeared to be unstable in JRS cells. These results indicate that co‐expression of the H205P and Y398Lfs*4 variants leads to a significant reduction in Asn synthesis and cellular growth. John Wiley & Sons, Inc. 2023-01-05 /pmc/articles/PMC9981421/ /pubmed/36873094 http://dx.doi.org/10.1002/jmd2.12356 Text en © 2022 The Authors. JIMD Reports published by John Wiley & Sons Ltd on behalf of SSIEM. https://creativecommons.org/licenses/by/4.0/This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research Reports
Staklinski, Stephen J.
Chang, Mario C.
Ahrens‐Nicklas, Rebecca C.
Kaur, Shagun
Stefanatos, Arianna K.
Dudenhausen, Elizabeth E.
Merritt, Matthew E.
Kilberg, Michael S.
Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines
title Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines
title_full Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines
title_fullStr Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines
title_full_unstemmed Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines
title_short Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines
title_sort characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines
topic Research Reports
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9981421/
https://www.ncbi.nlm.nih.gov/pubmed/36873094
http://dx.doi.org/10.1002/jmd2.12356
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