Translation of yeast GCN4 mRNA occurs by a reinitiation mechanism that is 
modulated by amino acid levels in the cell. Ribosomes which translate the first 
of four upstream open reading frames (uORFs) in the mRNA leader resume scanning 
and can reinitiate downstream. Under non-starvation conditions reinitiation 
occurs at one of the remaining three uORFs and GCN4 is repressed. Under 
starvation conditions, in contrast, ribosomes bypass the uORFs and reinitiate at 
GCN4 instead. The high frequency of reinitiation following uORF1 translation 
depends on an adequate distance to the next start codon and particular sequences 
surrounding the uORF1 stop codon. We present evidence that sequences 5' to uORF1 
also strongly enhance reinitiation. First, reinitiation was severely inhibited 
when uORF1 was transplanted into the position of uORF4, even though the native 
sequence environment of the uORF1 stop codon was maintained, and this effect 
could not be accounted for by the decreased uORF1-GCN4 spacing. Second, 
insertions and deletions in the leader preceding uORF1 greatly reduced 
reinitiation at GCN4. Sequences 5' to uORF1 may influence the probability of 
ribosome release following peptide termination at uORF1. Alternatively, they may 
facilitate rebinding of an initiation factor required for reinitiation prior to 
resumption of the scanning process.