Kozak sequences regulate gene expression in Trypanosoma brucei

The Kozak consensus sequence around the AUG start codon of mRNAs allows efficient cytosolic translation initiation in eukaryotes. It has not yet been investigated in the parasitic protozoan Trypanosoma brucei. mRNAs in T. brucei and other kinetoplastids are exclusively produced by polycistronic transcription. The levels of individual mRNAs can, therefore, not be regulated by transcription. Here, we show that in contrast to yeast and animals, no Kozak consensus sequence could be found in the total mRNA population of T. brucei or other kinetoplastids. However, when analyzing the subpopulation of constitutively expressed trypanosomal mRNAs encoding highly abundant proteins, a Kozak motif with a strong bias toward a +5C was detected. We tested how variants of the Kozak sequence influence the translation levels of a reporter protein. Using this in vivo approach, "weak" and "strong" Kozak sequences resulting in "low" and "high" translation levels could be defined. The "strong" sequences required a +5C and allowed initiation from CUG instead of AUG. We, therefore, suggest that in T. brucei, due to the lack of transcriptional control, Kozak sequences contribute to the regulation of protein levels. Moreover, we provide a new way to modulate protein abundance in transgenic trypanosomes in a predicted way.