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Intron retention resulting from a silent mutation in the VWF gene that structurally influences the 5' splice site.
Blood 2016; 128(17):2144-2152Blood

Abstract

Disease-associated silent mutations are considered to affect the accurate pre-messenger RNA (mRNA) splicing either by influencing regulatory elements, leading to exon skipping, or by creating a new cryptic splice site. This study describes a new molecular pathological mechanism by which a silent mutation inhibits splicing and leads to intron retention. We identified a heterozygous silent mutation, c.7464C>T, in exon 44 of the von Willebrand factor (VWF) gene in a family with type 1 von Willebrand disease. In vivo and ex vivo transcript analysis revealed an aberrantly spliced transcript, with intron 44 retained in the mRNA, implying disruption of the first catalytic step of splicing at the 5' splice site (5'ss). The abnormal transcript with the retained intronic region coded a truncated protein that lacked the carboxy-terminal end of the VWF protein. Confocal immunofluorescence characterizations of blood outgrowth endothelial cells derived from the patient confirmed the presence of the truncated protein by demonstrating accumulation of VWF in the endoplasmic reticulum. In silico pre-mRNA secondary and tertiary structure analysis revealed that this substitution, despite its distal position from the 5'ss (85 bp downstream), induces cis alterations in pre-mRNA structure that result in the formation of a stable hairpin at the 5'ss. This hairpin sequesters the 5'ss residues involved in U1 small nuclear RNA interactions, thereby inhibiting excision of the pre-mRNA intronic region. This study is the first to show the allosteric-like/far-reaching effect of an exonic variation on pre-mRNA splicing that is mediated by structural changes in the pre-mRNA.

Authors+Show Affiliations

Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany.Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany.Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany.Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany.Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany.Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany.Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany.

Pub Type(s)

Case Reports
Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

27543438

Citation

Yadegari, Hamideh, et al. "Intron Retention Resulting From a Silent Mutation in the VWF Gene That Structurally Influences the 5' Splice Site." Blood, vol. 128, no. 17, 2016, pp. 2144-2152.
Yadegari H, Biswas A, Akhter MS, et al. Intron retention resulting from a silent mutation in the VWF gene that structurally influences the 5' splice site. Blood. 2016;128(17):2144-2152.
Yadegari, H., Biswas, A., Akhter, M. S., Driesen, J., Ivaskevicius, V., Marquardt, N., & Oldenburg, J. (2016). Intron retention resulting from a silent mutation in the VWF gene that structurally influences the 5' splice site. Blood, 128(17), pp. 2144-2152.
Yadegari H, et al. Intron Retention Resulting From a Silent Mutation in the VWF Gene That Structurally Influences the 5' Splice Site. Blood. 2016 10 27;128(17):2144-2152. PubMed PMID: 27543438.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Intron retention resulting from a silent mutation in the VWF gene that structurally influences the 5' splice site. AU - Yadegari,Hamideh, AU - Biswas,Arijit, AU - Akhter,Mohammad Suhail, AU - Driesen,Julia, AU - Ivaskevicius,Vytautas, AU - Marquardt,Natascha, AU - Oldenburg,Johannes, Y1 - 2016/08/19/ PY - 2016/02/12/received PY - 2016/08/08/accepted PY - 2016/10/30/pubmed PY - 2017/7/29/medline PY - 2016/8/21/entrez SP - 2144 EP - 2152 JF - Blood JO - Blood VL - 128 IS - 17 N2 - Disease-associated silent mutations are considered to affect the accurate pre-messenger RNA (mRNA) splicing either by influencing regulatory elements, leading to exon skipping, or by creating a new cryptic splice site. This study describes a new molecular pathological mechanism by which a silent mutation inhibits splicing and leads to intron retention. We identified a heterozygous silent mutation, c.7464C>T, in exon 44 of the von Willebrand factor (VWF) gene in a family with type 1 von Willebrand disease. In vivo and ex vivo transcript analysis revealed an aberrantly spliced transcript, with intron 44 retained in the mRNA, implying disruption of the first catalytic step of splicing at the 5' splice site (5'ss). The abnormal transcript with the retained intronic region coded a truncated protein that lacked the carboxy-terminal end of the VWF protein. Confocal immunofluorescence characterizations of blood outgrowth endothelial cells derived from the patient confirmed the presence of the truncated protein by demonstrating accumulation of VWF in the endoplasmic reticulum. In silico pre-mRNA secondary and tertiary structure analysis revealed that this substitution, despite its distal position from the 5'ss (85 bp downstream), induces cis alterations in pre-mRNA structure that result in the formation of a stable hairpin at the 5'ss. This hairpin sequesters the 5'ss residues involved in U1 small nuclear RNA interactions, thereby inhibiting excision of the pre-mRNA intronic region. This study is the first to show the allosteric-like/far-reaching effect of an exonic variation on pre-mRNA splicing that is mediated by structural changes in the pre-mRNA. SN - 1528-0020 UR - https://www.unboundmedicine.com/medline/citation/27543438/Intron_retention_resulting_from_a_silent_mutation_in_the_VWF_gene_that_structurally_influences_the_5'_splice_site_ L2 - http://www.bloodjournal.org/cgi/pmidlookup?view=long&pmid=27543438 DB - PRIME DP - Unbound Medicine ER -