Evaluation of the role of ATP-binding cassette transporters as a defence mechanism against temephos in populations of Aedes aegypti

The role of ATP-binding cassette (ABC) transporters in the efflux of the insecticide, temephos, was assessed in the larvae of Aedes aegypti. Bioassays were conducted using mosquito populations that were either susceptible or resistant to temephos by exposure to insecticide alone or in combination with sublethal doses of the ABC transporter inhibitor, verapamil (30, 35 and 40 μM). The best result in the series was obtained with the addition of verapamil (40 μM), which led to a 2x increase in the toxicity of temephos, suggesting that ABC transporters may be partially involved in conferring resistance to the populations evaluated.

Down-regulation of ATP-binding cassette transporter G1 expression by unmethylated CpG oligodeoxynucleotides in RAW 264.7 macrophages

We have investigated the effect of various forms of phosphodiester cytidine-phosphate-guanosine oligodeoxynucleotides (CpG ODNs) on the production of pro-inflammatory cytokines and related genes in RAW 264.7 macrophages. Treatment with the CpG ODNs increased the expression of tumor necrosis factor alpha (TNF-alpha), IL-6, and inducible nitric oxide synthase but not interleukin-1beta (IL-1beta). We also investigated the effect of CpG ODNs on the expression of ATP-binding cassette transporter A1 (ABCA1) and G1 (ABCG1) genes which are known to facilitate cholesterol efflux from macrophages for anti-atherosclerosis. CpG 2006 significantly reduced the levels of ABCG1 mRNA as determined by real-time polymerase chain reaction, whereas ABCA1 mRNA level was not changed. Western blot analysis further confirmed the reduction of ABCG1 protein expression by CpG 2006. In addition, we also determined the protein level of peroxisome proliferator activated receptor gamma (PPARgamma), which is recognized as a transcriptional activator of ABC transporters, was also reduced by CpG 2006. Thus, these results suggest that ABCG1 is specifically down-regulated by CpG 2006 in a PPARgamma-dependent manner in macrophages.

Estimation of efflux mediated multi-drug resistance and its correlation with expression levels of two major efflux pumps in mycobacteria.

Multidrug resistance has been posing an increasing problem in the treatment of tuberculosis. Mutations in the genomic targets of drugs have been identified as the major mechanism behind this resistance. However, high degree of resistance in some isolates towards major drugs like rifampicin, isoniazid, ethambutol and streptomycin can not be explained solely on the basis of mutations. Besides this, certain other mechanisms like efflux pumps have also been considered as alternative mechanisms in the drug resistant isolates where there is no mutation and these mechanisms are specially important for drug resistance in non-tuberculous mycobacteria (NTM). In this study, we have estimated efflux pump mediated drug resistance in different mycobacterial species with the help of efflux pump inhibitors. All major anti-tuberculous drugs have been shown to be extruded by efflux pumps and the degree to which these drugs are extruded, vary in different mycobacterial species and isolates. The correlation of this resistance with functional activity of two major efflux pump genes pstB and Rv1258c was also assessed by reverse transcription PCR. Besides the significant role of these pumps observed, other efflux pumps, present in mycobacteria, may also be involved in drug resistance and need to be investigated.

Transporters in the Paracoccidioides brasiliensis transcriptome: insights on drug resistance

In the struggle for life, the capacity of microorganisms to synthesize and secrete toxic compounds (inhibiting competitors) plays an important role in successful survival of these species. This ability must come together with the capability of being unaffected by these same compounds. Several mechanisms are thought to avoid the toxic effects. One of them is toxin extrusion from the intracellular environment to the outside vicinity, using special transmembrane proteins, referred to as transporters. These proteins are also important for other reasons, since most of them are involved in nutrient uptake and cellular excretion. In cancer cells and in pathogens, and particularly in fungi, some of these proteins have been pointed out as responsible for an important phenotype known as multidrug resistance (MDR). In the present study, we tried to identify in the Paracoccidioides brasiliensis transcriptome, transporter-ortholog genes from the two major classes: ATP binding cassette and major facilitator superfamily transporter. We found 22 groups with good similarity with other fungal ATP binding cassette transporters, and four Paracoccidioides brasilienses assembled expressed sequence tags that probably code for major facilitator superfamily proteins. We also focused on fungicide resistance orthologs already characterized in other pathogenic fungi. We were able to find homologs to C. albicans CDR1, CDR2, and MDR1, Saccharomyces cerevisiae PDR5 and Aspergillus AtrF genes, all of them related to azole resistance. As current treatment for paracoccidioidomycosis mainly uses azole derivatives, the presence of these genes can be postulated to play a similar role in P. brasiliensis, warning us for the possibility of resistant isolate emergence.