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Analysis of the consequences of premature termination codons within factor VIII coding sequences
Author(s) -
David D.,
Santos I. M. A.,
Johnson K.,
Tuddenham E. G. D.,
McVey J. H.
Publication year - 2003
Publication title -
journal of thrombosis and haemostasis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.947
H-Index - 178
eISSN - 1538-7836
pISSN - 1538-7933
DOI - 10.1046/j.1538-7836.2003.00013.x
Subject(s) - nonsense mediated decay , messenger rna , nonsense mutation , transfection , microbiology and biotechnology , mutation , biology , stop codon , antibody , coding region , complementary dna , translation (biology) , rna splicing , genetics , gene , rna , missense mutation
Summary. Inhibitor antibody formation is a complication of factor VIII (FVIII) replacement therapy due to a failure to synthesize sufficient FVIII protein to induce immune tolerance. The incidence of nonsense mutations in inhibitor patients is high, however, this association is variable according to the position of the mutation. We have studied the effect of nonsense mutations on accumulation of FVIII mRNA, protein translation and secretion. Appropriately processed mRNA was detected in cells transfected with wild‐type R1966X and R2116X expression constructs and no evidence of nonsense‐mediated decay was observed. All constructs directed the translation of detectable intracellular FVIII antigen, however, secreted FVIII was detected only in conditioned media of cells transfected with wild‐type cDNA. We have also analyzed ectopic FVIII mRNA transcripts in the lymphocytes of six hemophilia A patients with nonsense mutations (Q139X, R583X, R1941X, R1966X and two unrelated patients with R2116X). FVIII mRNA was detectable in every case. In R1941X and R1966X only normally spliced transcripts were present. In Q139X, R583X and R2116X aberrantly spliced transcripts were observed with two distinct patterns in two individuals with the R2116X mutation. No correlation between mutation, transcript pattern and incidence of inhibitor development was apparent.