Molecular characterization of the hexose transporter gene in benznidazole resistant and susceptible populations of <it>Trypanosoma cruzi</it>

• Main Authors: dos Santos Paula F, Ruiz Jerônimo C, Soares Rodrigo P P, Moreira Douglas S, Rezende Antônio M, Folador Edson L, Oliveira Guilherme, Romanha Alvaro J, Murta Silvane MF
• Format: Article
• Language: English
• Published: BMC 2012-08-01
• Series: Parasites & Vectors
• Subjects: Trypanosoma cruzi; Drug resistance; Hexose transporters
• Online Access: http://www.parasitesandvectors.com/content/5/1/161

<p>Abstract</p> <p>Background</p> <p>Hexose transporters (HT) are membrane proteins involved in the uptake of energy-supplying glucose and other hexoses into the cell. Previous studies employing the Differential Display technique have shown that the transcription level of the HT gene from <it>T. cruzi</it> (<it>TcrHT</it>) is higher in an <it>in vitro-</it>induced benznidazole (BZ)-resistant population of the parasite (17 LER) than in its susceptible counterpart (17 WTS).</p> <p>Methods</p> <p>In the present study, <it>TcrHT</it> has been characterized in populations and strains of <it>T. cruzi</it> that are resistant or susceptible to BZ. We investigated the copy number and chromosomal location of the gene, the levels of <it>TcrHT</it> mRNA and of TcrHT activity, and the phylogenetic relationship between TcrHT and HTs from other organisms.</p> <p>Results</p> <p><it>In silico</it> analyses revealed that 15 sequences of the TcrHT gene are present in the <it>T. cruzi</it> genome, considering both CL Brener haplotypes. Southern blot analyses confirmed that the gene is present as a multicopy tandem array and indicated a nucleotide sequence polymorphism associated to <it>T. cruzi</it> group I or II. Karyotype analyses revealed that <it>TcrHT</it> is located in two chromosomal bands varying in size from 1.85 to 2.6 Mb depending on the strain of <it>T. cruzi</it>. The sequence of amino acids in the HT from <it>T. cruzi</it> is closely related to the HT sequences of <it>Leishmania</it> species according to phylogenetic analysis. Northern blot and quantitative real-time reverse transcriptase polymerase chain reaction analyses revealed that <it>TcrHT</it> transcripts are 2.6-fold higher in the resistant 17 LER population than in the susceptible 17 WTS. Interestingly, the hexose transporter activity was 40% lower in the 17 LER population than in all other <it>T. cruzi</it> samples analyzed. This phenotype was detected only in the <it>in vitro-</it>induced BZ resistant population, but not in the <it>in vivo</it>-selected or naturally BZ resistant <it>T. cruzi</it> samples. Sequencing analysis revealed that the amino acid sequences of the TcrHT from 17WTS and 17LER populations are identical. This result suggests that the difference in glucose transport between 17WTS and 17LER populations is not due to point mutations, but probably due to lower protein expression level.</p> <p>Conclusion</p> <p>The BZ resistant population 17 LER presents a decrease in glucose uptake in response to drug pressure.</p>