Myristoyl-CoA:protein N-myristoyltransferase depletion in trypanosomes causes avirulence and endocytic defects.
The enzyme myristoyl-CoA:protein N-myristoyltransferase (NMT) catalyses the co-translational covalent attachment of the fatty acid myristate to the N-terminus of target proteins. NMT is known to be essential for viability in Trypanosoma brucei and Leishmania major. Here we describe phenotypic analysis of T. brucei bloodstream form cells following knockdown of NMT expression by tetracycline-inducible RNA interference. Cell death occurs from 72h post-induction, with approximately 50% of cells displaying a defect in endocytic uptake by this time. The majority of these induced cells do not have an enlarged flagellar pocket typical of a block in endocytosis but vesicle accumulation around the flagellar pocket indicates a defect in vesicular progression following endocytic fusion. Induced parasites have a wild-type or slightly enlarged Golgi apparatus, unlike the phenotype of cells with reduced expression of a major N-myristoylated protein, ARL1. Critically we show that following NMT knockdown, T. brucei bloodstream form cells are unable to establish an infection in a mouse model, therefore providing further validation of this enzyme as a target for drug development.
Price
Helen P
HP
Centre for Immunology and Infection, Department of Biology, University of York, UK. hp502@york.ac.uk
Güther
M Lucia S
ML
Ferguson
Michael A J
MA
Smith
Deborah F
DF
D000217:Acyltransferases / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D004705:Endocytosis; D055785:Gene Knockdown Techniques; D051379:Mice; D008807:Mice, Inbred BALB C; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology; D014353:Trypanosomiasis, African / Q000469:parasitology*; D014774:Virulence
D015800:Protozoan Proteins; D000217:Acyltransferases; C050480:glycylpeptide N-tetradecanoyltransferase