Increased functional protein expression using nucleotide sequence features enriched in highly expressed genes in zebrafish.
Bottom Line: Zebrafish embryos are rapidly injected with calibrated doses of mRNA, enabling the effects of multiple sequence changes to be compared in vivo.These results suggested principles for increasing protein yield in zebrafish through biomolecular engineering.Rational gene design thus significantly boosts expression in zebrafish, and a similar approach will likely elevate expression in other animal models.
Affiliation: Program in Genomics of Differentiation, Eunice Kennedy Shriver National Institute of Child Health and Human Development, Bethesda, MD 20892, USA.Show MeSH
Mentions: Translation initiation is impaired when strong secondary structure for nucleotides −4 to +37 (relative to the AUG) reduces accessibility for the ribosome (11,61). In other parts of the transcript, local mRNA structure is not generally a major determinant of the rate of translation elongation since the ribosome processively destabilizes the mRNA (62,63). The mean free energy of the minimum energy structure (dG) at 28°C for zebrafish transcripts was 2 kcal/mol greater than for mouse or human transcripts, suggesting that nucleotide sequence has adapted to reduce secondary structure at the typical environmental temperatures that zebrafish inhabit (16.5–33°C (64); Figure 3a). Highly expressed genes showed a small but significant tendency for a more open mRNA structure than other genes (Figure 3a and b).
Affiliation: Program in Genomics of Differentiation, Eunice Kennedy Shriver National Institute of Child Health and Human Development, Bethesda, MD 20892, USA.