Abstract

The binding of thiamine pryrophosphate (TPP) to the aptamer of the THI4 riboswitch redirects expression platform alternative splicing to include an upstream open reading frame (uORF) in the transcript; the uORF precedes the THI4 codon sequence thereby interfering withTHI4 translation. There has been thorough study of the nucleotides and secondary structures of the THI4 aptamer, but less attention dedicated to residues in the expression platform. In particular, are the expression platform nucleotides active or passive in riboswitch regulated alternative splicing? Do nucleotypic characteristics of the expression platform, including size and GC content, influence regulation? Can the uORF be modified? Through the assembly of six different THI4 constructs and phenotypic analysis in C. reinhardtii this presentation addresses the abovementioned questions as well as highlights the implications for using the THI4 riboswitch as a synthetic biology tool.