Nucleic Acids Research, 2001, Vol. 29, No. 3 716-724
© 2001 Oxford University Press
An alteration in concatameric structure is associated with efficient segregation of plasmids in transfected Plasmodium falciparum parasites
Department of Microbiology and Immunology and the CRC for Vaccine Technology, The University of Melbourne, VIC 3010, Australia, 1Division of Parasitology, The National Institute for Medical Research, The Ridgeway, Mill Hill, London NW7 1AA, UK and 2The Walter and Eliza Hall Institute of Medical Research, PO Royal Melbourne Hospital, Melbourne, VIC 3050, Australia
Transfection of the human malaria parasite Plasmodium falciparum is currently performed with circularised plasmids that are maintained episomally in parasites under drug selection but which are rapidly lost when selection pressure is removed. In this paper, we show that in instances where gene targeting is not favoured, transfected plasmids can change to stably replicating forms (SRFs) that are maintained episomally in the absence of drug selection. SRF DNA is a large concatamer of the parental plasmid comprising at least nine plasmids arranged in a head-to-tail array. We show as well that the original unstable replicating forms (URFs) are also present as head-to-tail concatamers, but only comprise three plasmids. Limited digestion and 
irradiation experiments revealed that while URF concatamers are primarily circular, as expected, SRF concatamers form a more complex structure that includes extensive single-stranded DNA. No evidence of sequence rearrangement or additional sequence was detected in SRF DNA, including in transient replication experiments designed to select for more efficiently replicating plasmids. Surprisingly, these experiments revealed that the bacterial plasmid alone can replicate in parasites. Together, these results imply that transfected plasmids are required to form head-to-tail concatamers to be maintained in parasites and implicate both rolling-circle and recombination-dependent mechanisms in their replication.
* To whom correspondence should be addressed at: Department of Microbiology and Immunology, The University of Melbourne, VIC 3010, Australia. Tel: +61 3 8344 5705; Fax: +61 3 9347 1540; Email: crabb{at}wehi.edu.au
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  N. A. Tchurikov, O. V. Kretova, E. D. Moiseeva, and D. V. Sosin Evidence for RNA synthesis in the intergenic region between enhancer and promoter and its inhibition by insulators in Drosophila melanogaster Nucleic Acids Res., January 1, 2009; 37(1): 111 - 122. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. R. Sanders, L. M. Kats, D. R. Drew, R. A. O'Donnell, M. O'Neill, A. G. Maier, R. L. Coppel, and B. S. Crabb A Set of Glycosylphosphatidyl Inositol-Anchored Membrane Proteins of Plasmodium falciparum Is Refractory to Genetic Deletion Infect. Immun., July 1, 2006; 74(7): 4330 - 4338. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Meissner, E. Krejany, P. R. Gilson, T. F. de Koning-Ward, D. Soldati, and B. S. Crabb Tetracycline analogue-regulated transgene expression in Plasmodium falciparum blood stages using Toxoplasma gondii transactivators PNAS, February 22, 2005; 102(8): 2980 - 2985. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Boucher, F. McNicoll, M. Laverdiere, A. Rochette, M.-N. Chou, and B. Papadopoulou The ribosomal RNA gene promoter and adjacent cis-acting DNA sequences govern plasmid DNA partitioning and stable inheritance in the parasitic protozoan Leishmania Nucleic Acids Res., May 25, 2004; 32(9): 2925 - 2936. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. R. Drew, R. A. O'Donnell, B. J. Smith, and B. S. Crabb A Common Cross-species Function for the Double Epidermal Growth Factor-like Modules of the Highly Divergent Plasmodium Surface Proteins MSP-1 and MSP-8 J. Biol. Chem., May 7, 2004; 279(19): 20147 - 20153. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. L. Baldi, R. Good, M. T. Duraisingh, B. S. Crabb, and A. F. Cowman Identification and Disruption of the Gene Encoding the Third Member of the Low-Molecular-Mass Rhoptry Complex in Plasmodium falciparum Infect. Immun., September 1, 2002; 70(9): 5236 - 5245. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Z. Krnajski, T.-W. Gilberger, R. D. Walter, A. F. Cowman, and S. Muller Thioredoxin Reductase Is Essential for the Survival of Plasmodium falciparum Erythrocytic Stages J. Biol. Chem., July 12, 2002; 277(29): 25970 - 25975. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. H. Williamson, C. J. Janse, P. W. Moore, A. P. Waters, and P. R. Preiser Topology and replication of a nuclear episomal plasmid in the rodent malaria Plasmodium berghei Nucleic Acids Res., February 1, 2002; 30(3): 726 - 731. [Abstract] [Full Text] [PDF]
|
|