Predominance of six different hexanucleotide recoding signals 3' of read-through stop codons

Abstract

Redefinition of UAG, UAA and UGA to specify a standard amino acid occurs in response to recoding signals present in a minority of mRNAs. This 'read-through' is in competition with termination and is utilized for gene expression. One of the recoding signals known to stimulate read-through is a hexanucleotide sequence of the form CARYYA 3' adjacent to the stop codon. The present work finds that of the 91 unique viral sequences annotated as read-through, 90% had one of six of the 64 possible codons immediately 3' of the read-through stop codon. The relative efficiency of these read-through contexts in mammalian tissue culture cells has been determined using a dual luciferase fusion reporter. The relative importance of the identity of several individual nucleotides in the different hexanucleotides is complex.

Figure 1

Examination of non-randomness in read-through groups. Each data set represents the 82 nt region surrounding either the leaky (left column) or non-leaky (right column) stop codon for each of the six +1 triplet groups. For each data set, the x-axis represents the individual nucleotide position from the –19th to the +60th relative to the stop codon, and the y-axis is a measure of the non-randomness associated with each nucleotide position in the form of a c² value.

Figure 2

Read-through facilitated by p2luc constructs. Each of the 15 sequences listed on the x-axis were tested for their ability to facilitate read-through of Photinus luciferase using the fusion vector p2luc. Sequences 1, 4, 11, 12 and 14 represent exactly the +1 and +2 triplet sequences from groups 1, 3, 4, 2 and 2 respectively. The other 10 sequences were either substitutions of a single nucleotide position (sequences 2, 3 and 5–9) or a substitution of the stop codon (sequences 10, 13 and 15).