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Evidence for an alternate splicing in the thyroperoxidase messenger from patients with Graves' disease
E Zanelli1, M Henry, B Charvet
1Laboratoire de Biochimie Médicale, U.38 INSERM, Faculté de Médecine, Marseille, France.
Abstract:
An initial lambda gt 11 cDNA library constructed from a human Graves' patient thyroid was screened with an immunopurified rabbit anti-human thyroperoxidase (hTPO) polyclonal antibody. A 869 bp clone was obtained. It presents a 130 bp deletion as compared to the published sequence and a 77 bp insertion in the 3' non-coding region. Screening of a pUC cDNA library from another Graves' patient thyroid exhibited the same 130 bp deletion in two other cDNA clones. PCR analysis of mRNA transcripts confirmed the presence of the two messengers in two other Graves' thyroid tissues. In all the cases, this new spliced mRNA species represents between 40% and 50% of the total hTPO mRNAs. With respect to the structure of the hTPO gene, the present deletion suggests an alternate splicing of exon 16. The juxtaposition of exon 17 to exon 15 encoding the transmembrane domain leads to a shift in the reading frame. By the use of a different stop codon, the spliced mRNA generates a modified 56 - COOH terminal aminoacids (aa) sequence.
Insights
Researchers identified a novel spliced mRNA variant in Graves' disease patients, affecting human thyroperoxidase (hTPO) production. This alternative splicing results in a modified protein, potentially impacting thyroid function in autoimmune thyroid disease.
Area of Science:
- Molecular Biology
- Endocrinology
- Genetics
Background:
- Graves' disease is an autoimmune disorder affecting the thyroid gland.
- Human thyroperoxidase (hTPO) is a key enzyme in thyroid hormone synthesis.
- Aberrant gene expression can contribute to thyroid dysfunction.
Purpose of the Study:
- To investigate potential molecular alterations in human thyroperoxidase (hTPO) in Graves' disease patients.
- To identify and characterize novel hTPO mRNA variants.
Main Methods:
- Screening of human Graves' patient thyroid cDNA libraries (lambda gt 11 and pUC).
- Characterization of cDNA clones using sequencing and PCR analysis of mRNA transcripts.
- Analysis of hTPO gene structure and potential alternative splicing events.
Main Results:
- A novel 869 bp hTPO cDNA clone with a 130 bp deletion and a 77 bp insertion was identified.
- The 130 bp deletion was confirmed in other cDNA clones and mRNA transcripts from Graves' thyroid tissues.
- This alternatively spliced mRNA species constitutes 40-50% of total hTPO mRNAs in affected patients.
- The deletion results from alternative splicing of exon 16, leading to a frame shift and modified C-terminal amino acid sequence.
Conclusions:
- A significant proportion of hTPO mRNA in Graves' disease patients exhibits alternative splicing.
- This splicing event generates a modified hTPO protein with altered C-terminal sequence.
- The functional implications of this novel hTPO variant in Graves' disease pathogenesis warrant further investigation.