Abstract
BackgroundChromosome 9 of Trypanosoma brucei contains two closely spaced, very similar open reading frames for cyclic nucleotide specific phosphodiesterases TbrPDEB1 and TbrPDEB2. They are separated by 2379 bp, and both code for phosphodiesterases with two GAF domains in their N-terminal moieties and a catalytic domain at the C-terminus.Methods and FindingsThe current study reveals that in the Lister427 strain of T. brucei, these two genes have undergone gene conversion, replacing the coding region for the GAF-A domain of TbrPDEB2 by the corresponding region of the upstream gene TbrPDEB1. As a consequence, these strains express two slightly different versions of TbrPDEB2. TbrPDEB2a represents the wild-type phosphodiesterase, while TbrPDEB2b represents the product of the converted gene. Earlier work on the subcellular localization of TbrPDEB1 and TbrPDEB2 had demonstrated that TbrPDEB1 is predominantly located in the flagellum, whereas TbrPDEB2 partially locates to the flagellum but largely remains in the cell body. The current findings raised the question of whether this dual localization of TbrPDEB2 may reflect the two alleles. To resolve this, TbrPDEB2 of strain STIB247 that is homozygous for TbrPDEB2a was tagged in situ, and its intracellular localization was analyzed.ConclusionsThe results obtained were very similar to those found earlier with Lister427, indicating that the dual localization of TbrPDEB2 reflects its true function and is not simply due to the presence of the two different alleles. Notably, the gene conversion event is unique for the Lister427 strain and all its derivatives. Based on this finding, a convenient PCR test has been developed that allows the stringent discrimination between Lister-derived strains that are common in many laboratories and other isolates. The technique is likely very useful to resolve questions about potential mix-ups of precious field isolates with the ubiquitous Lister strain.
Highlights
Cyclic nucleotide-specific phosphodiesterases (PDEs) are crucial players in cyclic nucleotide signalling of eukaryotic cells
One allele corresponded to that present in the T. brucei database (TbrPDEB2a)
In the second allele (TbrPDEB2b), a part of the sequence that codes for the GAF-A domain was replaced by the corresponding sequence from TbrPDEB1 (Fig. 1)
Summary
Cyclic nucleotide-specific phosphodiesterases (PDEs) are crucial players in cyclic nucleotide signalling of eukaryotic cells. The genomes of all kinetoplastid parasites, including Trypanosoma brucei, contain similar sets of genes that code for cyclic nucleotide-specific phosphodiesterases (PDEs; [2,5]). In T. brucei, these two genes, TbrPDEB1 (Tb09.160.3590) and TbrPDEB2 (Tb09.160.3630), are located on chromosome 9, and their open reading frames are separated by 2379 bp They code for very similar proteins of 930 and 925 amino acids, respectively. Chromosome 9 of Trypanosoma brucei contains two closely spaced, very similar open reading frames for cyclic nucleotide specific phosphodiesterases TbrPDEB1 and TbrPDEB2. They are separated by 2379 bp, and both code for phosphodiesterases with two GAF domains in their N-terminal moieties and a catalytic domain at the C-terminus
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