RNA polymerase I promoter and splice acceptor site recognition affect gene expression in non-pathogenic Leishmania species.
Leishmania (Sauroleishmania) tarentolae has biotechnological potential for use as live vaccine against visceral leishmaniasis and as a system for the over expression of eukaryotic proteins that possess accurate post-translational modifications. For both purposes, new systems for protein expression in this non-pathogenic protozoan are necessary. The ribosomal RNA promoter proved to be a stronger transcription driver since its use yielded increased levels of recombinant protein in organisms of both genera Trypanosoma or Leishmania. We have evaluated heterologous expression systems using vectors with two different polypyrimidine tracts in the splice acceptor site by measuring a reporter gene transcribed from L. tarentolae RNA polymerase I promoter. Our data indicate that the efficiency of chloramphenicol acetyl transferase expression changed drastically with homologous or heterologous sequences, depending on the polypyrimidine tract used in the construct and differences in size and/or distance from the AG dinucleotide. In relation to the promoter sequence the reporter expression was higher in heterologous lizard-infecting species than in the homologous L. tarentolae or in the mammalian-infecting L. (Leishmania) amazonensis.
Orlando
Tereza Cristina
TC
Departamento de Fisiologia, Instituto de Biociências, São Paulo, SP, 05508-900, Brazil.
Mayer
Mário Gustavo
MG
Campbell
David A
DA
Sturm
Nancy R
NR
Floeter-Winter
Lucile Maria
LM
D000818:Animals; D015870:Gene Expression; D007891:Leishmania / Q000145:classification / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012318:RNA Polymerase I / Q000235:genetics*; D022821:RNA Splice Sites / Q000235:genetics*
D022821:RNA Splice Sites; D012318:RNA Polymerase I