Trypanosoma brucei glycosomal glyceraldehyde-3-phosphate dehydrogenase genes are stage-regulated at the transcriptional level

Abstract

Regions 5' of the glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) gene from Trypanosoma brucei were tested for their ability to promote chloramphenicol acetyl-transferase (CAT) expression on reintroduction by electroporation into the parasite. Deletion analysis mapped the gGAPDH promoter to within 403 nts of the start of translation. A transcription initiation site was mapped at around -190 nts from the ATG start codon by RNase protection and by primer extension. The higher expression of gGAPDH in bloodstream T. brucei, compared to procyclic (insect) forms, was largely attributed to differences in promoter activity. The gGAPDH promoter gave rise to relatively high CAT signals upon transfection into bloodstream T. brucei and relatively low signals in procyclic T. brucei, compared with levels resulting from transfection with the procyclic acidic repetitive protein (PARP) promoter. In addition, RNase protection data showed a higher level of gGAPDH primary transcripts in bloodstream. T. brucei. The PARP mini-exon addition region abolished transient CAT expression directed by either the gGAPDH or PARP promoters in bloodstream T. brucei implying that transplicing can be a point of stage-specific gene regulation.