Expression of human nPTB is limited by extreme suboptimal codon content.
BACKGROUND
The frequency of synonymous codon usage varies widely between organisms. Suboptimal codon content limits expression of viral, experimental or therapeutic heterologous proteins due to limiting cognate tRNAs. Codon content is therefore often adjusted to match codon bias of the host organism. Codon content also varies between genes within individual mammalian species. However, little attention has been paid to the consequences of codon content upon translation of host proteins.


RESULTS
In comparing the splicing repressor activities of transfected human PTB and its two tissue-restricted paralogs-nPTB and ROD1-we found that the three proteins were expressed at widely varying levels. nPTB was expressed at 1-3% the level of PTB despite similar levels of mRNA expression and 74% amino acid identity. The low nPTB expression was due to the high proportion of codons with A or U at the third codon position, which are suboptimal in human mRNAs. Optimization of the nPTB codon content, akin to the "humanization" of foreign ORFs, allowed efficient translation in vivo and in vitro to levels comparable with PTB. We were then able to demonstrate that all three proteins act as splicing repressors.


CONCLUSIONS
Our results provide a striking illustration of the importance of mRNA codon content in determining levels of protein expression, even within cells of the natural host species.
Robinson
Fiona
F
Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom.
Jackson
Richard J
RJ
Smith
Christopher W J
CW
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003062:Codon; D017931:DNA Primers; D015870:Gene Expression; D006801:Humans; D051379:Mice; D016366:Open Reading Frames; D038941:Polypyrimidine Tract-Binding Protein / Q000235:genetics*; D012313:RNA / Q000302:isolation & purification
D003062:Codon; D017931:DNA Primers; D038941:Polypyrimidine Tract-Binding Protein; D012313:RNA