Transcripts of a heterologous gene encoding mite allergen Der f 7 are stabilized by codon optimization in Aspergillus oryzae

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• 作者：Mizuki Tanaka (1)
  Masafumi Tokuoka (1) (2)
  Takahiro Shintani (1)
  Katsuya Gomi (1)
  
• 关键词：mRNA stability ; Aspergillus oryzae ; Codon optimization ; Heterologous gene expression ; 1 ; 10 ; Phenanthroline ; Nonstop mRNA decay
• 刊名：Applied Microbiology and Biotechnology
• 出版年：2012
• 出版时间：December 2012
• 年：2012
• 卷：96
• 期：5
• 页码：1275-1282
• 全文大小：385KB
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• 作者单位：Mizuki Tanaka (1)
  Masafumi Tokuoka (1) (2)
  Takahiro Shintani (1)
  Katsuya Gomi (1)
  
  1. Laboratory of Bioindustrial Genomics, Department of Bioindustrial Informatics and Genomics, Graduate School of Agricultural Science, Tohoku University, 1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai, 981-8555, Japan
  2. National Research Institute of Brewing, 3-7-1 Kagamiyama, Higashi-Hiroshima, Hiroshima, 739-0046, Japan
  
• ISSN：1432-0614

文摘

We have previously demonstrated that transcripts of an AT-biased heterologous gene encoding mite allergen Der f 7 from Dermatophagoides farinae were polyadenylated prematurely within the coding region when native cDNA was expressed in Aspergillus oryzae, and that this premature polyadenylation was prevented by the codon optimization of the Der f 7 gene, resulting in increased steady-state mRNA levels. In this study, we tested the stability of transcription products derived from expression constructs of the native and codon-optimized Der f 7 gene in A. oryzae using 1,10-phenanthroline as a transcription inhibitor. Transcription products of native Der f 7 cDNA fused to the A. oryzae glucoamylase gene (glaA) were rapidly degraded; the half-life of the mRNA was approximately 13?min. However, the half-life of codon-optimized Der f 7 mRNA fused to glaA was approximately 43?min, which was highly similar to that of endogenous glaA mRNA. These results indicate that Der f 7 mRNA is significantly stabilized by codon optimization. In addition, Der f 7 mRNA was stabilized by the codon optimization of only the 3-half region, where premature polyadenylation sites were exclusively situated; the half-life of the chimeric Der f 7 mRNA was approximately 39?min. This suggested that destabilization of native Der f 7 mRNA is mainly triggered by premature polyadenylation within the coding region. To the best of our knowledge, this is the first report to provide experimental evidence that heterologous mRNA is significantly stabilized by codon optimization in eukaryotic cells.