DNA Sequence Changes Resulting from Codon Optimization Affect Gene Expression in Pichia pastoris by Altering...

Area of Science:

• Synthetic Biology
• Molecular Biology
• Gene Expression Regulation

Background:

• Codon optimization is a common method to increase protein production.
• Unexpected transcriptional repression can occur post-optimization, even with unchanged amino acid sequences.

Purpose of the Study:

• Investigate the mechanisms behind codon optimization-induced transcriptional attenuation.
• Analyze the impact of codon optimization on chromatin accessibility and gene expression in *P. pastoris*.

Main Methods:

• Analyzed codon optimization effects on *0432* and *Fluc* genes in *P. pastoris*.
• Utilized histone H3 chromatin immunoprecipitation (ChIP) and DNase I hypersensitivity assays.
• Assessed gene expression with different promoters (P, P, P).

Main Results:

• Codon optimization led to significant mRNA reduction in both analyzed genes.
• Repressed genes showed increased nucleosome occupancy and decreased chromatin accessibility.
• Transcriptional attenuation and chromatin changes persisted irrespective of promoter strength.

Conclusions:

• Codon optimization can influence transcription via chromatin accessibility, not just translation.
• Synthetic gene design must consider chromatin-level effects to prevent transcriptional silencing.
• Integrating chromatin insights is crucial for optimizing gene expression outcomes.