Down regulation of S-adenosyl-L-methionine decarboxylase activity of Trypanosoma brucei during transition from long slender to short stumpy-like forms in axenic culture.
Long slender trypanosomes, isolated from infected mouse blood or from cryopreserved stabilates, respectively, were unable to grow in conditioned media (cMEM), prepared from the declining phase of axenic bloodstream form cultures. Additionally, mixtures of fresh medium and cMEM led to decreased growth rates and, in accordance to the amount of cMEM used, to a decreased S-adenosyl-L-methionine decarboxylase (Ado-MetDC; E.C. 4.1.1.50) activity. Since addition of polyamines could not overcome the process of transition from dividing to non-dividing cells and the intracellular S-adenosyl-L-methionine (AdoMet), ornithine and putrescine concentrations seemed unaltered during the course of cultivation, we questioned if polyamine metabolism is involved in this transition process. Activities of two key enzymes of polyamine metabolism, AdoMetDC and ornithine decarboxylase (ODC; E.C. 4.1.1.17) were therefore monitored during different growth stages. Our results revealed a specific activity of 44 pmol min-1 mg protein-1 for AdoMetDC and a KM of 10 mu M for AdoMet. Methylglyoxal bis(guanylhydrazone) showed a Ki of 6 mu M. The constant activity of the enzyme during a 7 h time-course in the presence of cycloheximide indicates a t1/2 of more than 7 h for the trypanosomal enzyme. Enzyme activity in trypanosomes isolated from infected laboratory animals and from logarithmic phase bloodstream or procyclic form cultures was high according to a high dividing rate, whereas enzyme activity in parasites isolated from the stationary phase of bloodstream from culture was negligible. In these cultures, AdoMetDC activity decreased with a t1/2 of 7 h during transition from long slender to short stumpy-like forms as soon as the stationary phase was reached. ODC activity was high (approximately 300 pmol min-1 mg protein-1) in dividing trypanosomes isolated from infected animals as well as from logarithmic phase bloodstream or procyclic form cultures and decreased also during transition with a t1/2 of 10 h.
Physiologisch-chemisches Institut der Universitat, Tubingen, Germany.
P M
PM
Selzer
F
F
Hesse
B
B
Hamm-Kunzelmann
K
K
Muhlstadt
H
H
Echner
M
M
Duszenko
D012437:Adenosylmethionine Decarboxylase / Q000235:genetics*; D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D046508:Culture Techniques; D015536:Down-Regulation; D005856:Germ-Free Life; D016015:Logistic Models; D009955:Ornithine Decarboxylase / Q000378:metabolism; D014176:Protein Biosynthesis; D014346:Trypanosoma brucei brucei / Q000201:enzymology / Q000235:genetics*
D009955:Ornithine Decarboxylase; D012437:Adenosylmethionine Decarboxylase