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Growth-based Determination and Biochemical Confirmation of Genetic Requirements for Protein Degradation in Saccharomyces cerevisiae
Published on: February 16, 2015
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Rationally designed, heterologous S. cerevisiae transcripts expose novel expression determinants.
Tuval Ben-Yehezkel1,2,3,4, Shimshi Atar1,4, Hadas Zur1
1a Department of Biomedical Engineering ; Tel-Aviv University ; Tel-Aviv , Israel.
RNA Biology
|July 16, 2015
Summary
Discovering how RNA transcript features influence protein levels is key. This study reveals silent mutations in unexplored transcript regions significantly alter protein expression in eukaryotes, offering new gene expression engineering rules.
Area of Science:
- Molecular Biology
- Synthetic Biology
- Genetics
Background:
- Deducing causal links between RNA features and protein levels from endogenous data is challenging.
- Previous studies on heterologous gene expression focused on 5' regions and prokaryotes.
Purpose of the Study:
- To systematically investigate the impact of various transcript regions on gene expression using synthetic biology.
- To uncover novel causal relationships between transcript features and protein levels in natural expression regimes.
Main Methods:
- Designed and analyzed 383 gene variants of the HRSVgp04 gene ORF in S. cerevisiae.
- Employed a synthetic biology approach to differentiate effects of transcript regions up to 240 nucleotides into the ORF.
- Investigated correlations between protein levels, folding energy, and codon decoding times.
Main Results:
- Silent mutations in the 5' untranslated region (UTR) caused up to 15-fold changes in protein levels.
- Synonymous mutations downstream (>120 nucleotides) modulated protein levels by 160%-300%.
- Protein abundance correlated with folding energy and mean relative codon decoding times (r=0.62).
Conclusions:
- Selection for folding strength in the ORF relates to translation regulation.
- Translation elongation and tRNA adaptation causally modify protein levels.
- Provides insights into transcript evolution, gene expression regulation, and synthetic gene engineering.
