A PROMOTER DIRECTING ALPHA-AMANITIN-SENSITIVE TRANSCRIPTION OF GARP, THE MAJOR SURFACE-ANTIGEN OF INSECT STAGE TRYPANOSOMA-CONGOLENSE

The major surface antigen of procyclic and epimastigote forms of Trypa nosoma congolense in the tsetse fly is GARP (glutamic acid/alanine-ric h protein), which is thought to be the analogue of procyclin/PARP in T rypanosoma brucei, We have studied two T.congolense GARP loci (the 4.3 and 4.4 loci) whose transcription is alpha-amanitin sensitive, Whilst a transcriptional gap 5' of the first GARP gene in the cloned region of the 4.4 locus could not be detected, such a gap was present in the 5' flank of the first GARP gene in the 4.3 locus, We have located a GA RP transcription start site and, using reporter gene constructs contai ning a putative GARP promoter region in transient transfection studies , we have demonstrated promoter activity for the test region in T.cong olense, There are species-specific differences in sequences regulating expression of the two major surface antigens, GARP and procyclin/PARP : the GARP promoter is inactive in T.brucei while the procyclin/PARP p romoter is inactive in T.congolense. We have defined the splice accept or site for the 4.3 GARP gene by sequencing and by 5' RT-PCR and demon strated microheterogeneity in GARP polyadenylation by 3' RT-PCR, It ap pears that some GARP and procyclin/PARP RNA processing signals, althou gh similar, are also species-specific.