title: Trypanosoma brucei: in vitro slender-to-stumpy differentiation of culture-adapted, monomorphic bloodstream forms.
abstract: Pleomorphic Trypanosoma brucei strains are characterized by their ability to differentiate from replicating long slender forms into non-dividing short stumpy forms in the mammalian host. The differentiation process can be efficiently induced in vitro by treatment with the membrane-permeable cAMP derivative 8-(4-chlorophenylthio)-cAMP (pCPTcAMP). In contrast, monomorphic T. brucei strains do not differentiate to stumpy forms in the host. Here, we show that exposure of monomorphic, culture-adapted T. brucei bloodstream forms to pCPTcAMP allowed their subsequent differentiation into short stumpy forms. The stumpy nature of pCPTcAMP-treated parasites was confirmed by (1) morphological change, (2) inhibition of growth and DNA synthesis, (3) cell cycle arrest in the G(1)/G(0) phase, (4) expression of NADH diaphorase activity and dihydrolipoamide dehydrogenase, (5) disappearance of the small subunit of ribonucleotide reductase, (6) up-regulation of the major lysosomal membrane protein, and (7) efficient transformation into replicating procyclic insect forms after induction with citrate/cis-aconitate. Our results indicate that the inability of monomorphic T. brucei bloodstream forms to differentiate into short stumpy forms in the host may be due to a failure in the signalling pathway rather than in the differentiation process itself. Treatment of monomorphic bloodstream trypanosomes with pCPTcAMP could be a useful method for identifying the genes involved in the slender-to-stumpy differentiation process.
authors.affiliation: Abteilung Parasitologie, Hygiene-Institut der Ruprecht-Karls Universität, Im Neuenheimer Feld, Heidelberg, Germany.
authors.forename: Tanja
authors.initials: T
authors.lastname: Breidbach
authors.forename: Elise
authors.initials: E
authors.lastname: Ngazoa
authors.forename: Dietmar
authors.initials: D
authors.lastname: Steverding
mesh_terms: D000818:Animals; D002453:Cell Cycle / Q000187:drug effects; D000242:Cyclic AMP / Q000031:analogs & derivatives* / Q000494:pharmacology*; D008058:Dihydrolipoamide Dehydrogenase / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D008018:Life Cycle Stages / Q000187:drug effects; D012264:Ribonucleotide Reductases / Q000037:antagonists & inhibitors / Q000378:metabolism; D013873:Thionucleotides / Q000494:pharmacology*; D014346:Trypanosoma brucei brucei / Q000187:drug effects / Q000201:enzymology / Q000254:growth & development*
chemical_list: D004791:Enzyme Inhibitors; D013873:Thionucleotides; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D000242:Cyclic AMP; D012264:Ribonucleotide Reductases; D008058:Dihydrolipoamide Dehydrogenase