Functional Studies on Novel RET Mutations and Their Implications for Genetic Counseling for Hirschsprung Disease
Hirschsprung disease (HSCR) is a genetic disorder characterized by the absence of ganglion cells in the gut. RET is considered to be the main susceptibility gene. In our previous screening of 83 HSCR patients, targeted exome sequencing identified nine rare variants of RET, most of which were new dis...
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Frontiers Media S.A.
2019-10-01
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| Series: | Frontiers in Genetics |
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| Online Access: | https://www.frontiersin.org/article/10.3389/fgene.2019.00924/full |
| _version_ | 1818328271153004544 |
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| author | Hui Wang Qi Li Zhen Zhang Ping Xiao Long Li Qian Jiang |
| author_facet | Hui Wang Qi Li Zhen Zhang Ping Xiao Long Li Qian Jiang |
| author_sort | Hui Wang |
| collection | DOAJ |
| description | Hirschsprung disease (HSCR) is a genetic disorder characterized by the absence of ganglion cells in the gut. RET is considered to be the main susceptibility gene. In our previous screening of 83 HSCR patients, targeted exome sequencing identified nine rare variants of RET, most of which were new discoveries. Here, we performed in vitro arrays with functional studies to investigate their effects. Two variants (p.R77C and p.R67insL) were demonstrated to disrupt the glycosylation of RET and affect its subcellular localization. Three nonsense mutations (p.W85X, p.E252X, and p.Y263X) could not produce detectable RET full-length protein, and the other three mutations (p.R770X, p.Q860X, and p.V778Afs*1) were translated into truncated proteins of predicted sizes. One canonical splice acceptor site mutation (c.2802-2 A > G) was verified to affect gene regulation through aberrant splicing. In addition, we explored the effects of read-through reagents on RET nonsense mutations and showed that G418 significantly increased the full-length RET protein expression of p.Y263X in a dose-dependent manner, together with a mild recovery of p-ERK and p-STAT3. Our data provide a functional analysis of novel RET mutations and suggest that all of the rare variants detected from patients with clinically severe HSCR are indeed pathogenic. Thus, our findings have implications for proper genetic counseling. |
| first_indexed | 2024-12-13T12:29:30Z |
| format | Article |
| id | doaj.art-8415ea00660c444d95e80f587ce616da |
| institution | Directory Open Access Journal |
| issn | 1664-8021 |
| language | English |
| last_indexed | 2024-12-13T12:29:30Z |
| publishDate | 2019-10-01 |
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| spelling | doaj.art-8415ea00660c444d95e80f587ce616da2022-12-21T23:46:04ZengFrontiers Media S.A.Frontiers in Genetics1664-80212019-10-011010.3389/fgene.2019.00924456249Functional Studies on Novel RET Mutations and Their Implications for Genetic Counseling for Hirschsprung DiseaseHui Wang0Qi Li1Zhen Zhang2Ping Xiao3Long Li4Qian Jiang5Department of Medical Genetics, Beijing Municipal Key Laboratory of Child Development and Nutriomics, Capital Institute of Pediatrics, Beijing, ChinaDepartment of General Surgery, Capital Institute of Pediatrics Affiliated Children’s Hospital, Beijing, ChinaDepartment of General Surgery, Capital Institute of Pediatrics Affiliated Children’s Hospital, Beijing, ChinaDepartment of Pathology, Capital Institute of Pediatrics Affiliated Children’s Hospital, Beijing, ChinaDepartment of General Surgery, Capital Institute of Pediatrics Affiliated Children’s Hospital, Beijing, ChinaDepartment of Medical Genetics, Beijing Municipal Key Laboratory of Child Development and Nutriomics, Capital Institute of Pediatrics, Beijing, ChinaHirschsprung disease (HSCR) is a genetic disorder characterized by the absence of ganglion cells in the gut. RET is considered to be the main susceptibility gene. In our previous screening of 83 HSCR patients, targeted exome sequencing identified nine rare variants of RET, most of which were new discoveries. Here, we performed in vitro arrays with functional studies to investigate their effects. Two variants (p.R77C and p.R67insL) were demonstrated to disrupt the glycosylation of RET and affect its subcellular localization. Three nonsense mutations (p.W85X, p.E252X, and p.Y263X) could not produce detectable RET full-length protein, and the other three mutations (p.R770X, p.Q860X, and p.V778Afs*1) were translated into truncated proteins of predicted sizes. One canonical splice acceptor site mutation (c.2802-2 A > G) was verified to affect gene regulation through aberrant splicing. In addition, we explored the effects of read-through reagents on RET nonsense mutations and showed that G418 significantly increased the full-length RET protein expression of p.Y263X in a dose-dependent manner, together with a mild recovery of p-ERK and p-STAT3. Our data provide a functional analysis of novel RET mutations and suggest that all of the rare variants detected from patients with clinically severe HSCR are indeed pathogenic. Thus, our findings have implications for proper genetic counseling.https://www.frontiersin.org/article/10.3389/fgene.2019.00924/fullHirschsprung diseasefunctional analysisRETsplice site mutationpremature termination codon |
| spellingShingle | Hui Wang Qi Li Zhen Zhang Ping Xiao Long Li Qian Jiang Functional Studies on Novel RET Mutations and Their Implications for Genetic Counseling for Hirschsprung Disease Frontiers in Genetics Hirschsprung disease functional analysis RET splice site mutation premature termination codon |
| title | Functional Studies on Novel RET Mutations and Their Implications for Genetic Counseling for Hirschsprung Disease |
| title_full | Functional Studies on Novel RET Mutations and Their Implications for Genetic Counseling for Hirschsprung Disease |
| title_fullStr | Functional Studies on Novel RET Mutations and Their Implications for Genetic Counseling for Hirschsprung Disease |
| title_full_unstemmed | Functional Studies on Novel RET Mutations and Their Implications for Genetic Counseling for Hirschsprung Disease |
| title_short | Functional Studies on Novel RET Mutations and Their Implications for Genetic Counseling for Hirschsprung Disease |
| title_sort | functional studies on novel ret mutations and their implications for genetic counseling for hirschsprung disease |
| topic | Hirschsprung disease functional analysis RET splice site mutation premature termination codon |
| url | https://www.frontiersin.org/article/10.3389/fgene.2019.00924/full |
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