Authors
J Edwards-Smallbone1; C J Merrick1; 1 Keele University Discussion
Telomeres are DNA-protein complexes at chromosome ends which serve to protect them from recognition by DNA repair machinery, end-to-end fusions, and sequence loss. Telomeric DNA from many species is stabilized by a multi-component complex which contains single- and double-stranded DNA binding proteins. However, no dedicated telomere-binding proteins apart from telomerase have yet been characterised in Plasmodium falciparum. We identified a homologue of the human telomere binding protein TRF1 by sequence and predicted structural similarity. We then attempted to manipulate its expression using a variety of established techniques including gene disruption by single and double-crossover recombination, over-expression of a DD-tagged version and glmS ribozyme-mediated knockdown. No genomic integration of constructs or reliable manipulation of gene expression was achieved, since in all cases the only surviving parasites carried mutated constructs. This occurred even where targeting strategies would not affect, or would regenerate an intact version of the endogenous locus, suggesting that the locus may not be targetable. Expression of the gene was confirmed by end-point PCR from cDNA and by Western blotting using antisera raised against PfTRF1 peptides. mRNA and protein were detectable only in gametocytes, in broad agreement with genome-wide studies showing low expression levels in blood-stage parasites, and subsequent upregulation in gametocytes. 
