Premium
Ribosome readthrough accounts for secreted full‐length factor IX in hemophilia B patients with nonsense mutations
Author(s) -
Pinotti Mirko,
Caruso Pierpaolo,
Canella Alessandro,
Campioni Matteo,
Tagariello Giuseppe,
Castaman Giancarlo,
Giacomelli Sofia,
Belvini Donata,
Bernardi Francesco
Publication year - 2012
Publication title -
human mutation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 162
eISSN - 1098-1004
pISSN - 1059-7794
DOI - 10.1002/humu.22120
Subject(s) - nonsense mutation , ribosome , biology , microbiology and biotechnology , haemophilia b , blot , context (archaeology) , factor ix , mutation , gene , recombinant dna , messenger rna , nonsense mediated decay , genetics , translation (biology) , nonsense , rna , missense mutation , haemophilia a , haemophilia , paleontology , rna splicing
We investigated the spontaneous ribosome readthrough, virtually unexplored in genes encoding secreted proteins, over coagulation F9 nonsense mutations. Expression of recombinant factor IX (FIX) in eukaryotic cells demonstrated appreciable levels of secreted FIX molecules for the mutations p.R162* (5 ± 0.3% of rFIX‐wt antigen levels), p.R294* (3.1 ± 1.1%) and p.R298* (2.5 ± 0.7%), but not for the p.L103*. Western blotting revealed a large proportion of truncated molecules, which correlated with small amounts of full‐length FIX (rFIX‐162*, ∼0.5%; rFIX‐294*; and rFIX‐298*, ∼0.2%). Western blotting of plasma from FIX deficient (Hemophilia B) patients revealed traces of full‐length FIX for the p.R294* and p.R298* mutations, but not for the p.L103* mutation that triggered major FIX mRNA decay. The detection of full‐length proteins has clinical implication, particularly for post‐therapeutic immunological complications in Hemophilia. Data in patients' plasma and in vitro, obtained in the proper protein context, support a ribosome readthrough gradient, consistent with its predicted determinants of efficiency. Hum Mutat 33:1373–1376, 2012. © 2012 Wiley Periodicals, Inc.