E-GEOD-41750 - Comparative Genomic Hybridization of Leishmania major SbIII resistant mutants and L. major WT

Status
Released on 1 May 2013, last updated on 7 May 2013
Organism
Leishmania major
Samples (12)
Array (1)
Protocols (7)
Description
SbIII clonal mutants and an isogenic WT clonal line. Genomic DNA from clonal WT or mutants were digested and hybridized to whole genome DNA microarrays. Antimonials are still the mainstay of treatment against Leishmaniasis but in the past decade resistance has been a severe threat. We carried out short read next generation sequencing (NGS) and comparative genomic hybridization (CGH) of three independent Leishmania major antimony resistant mutants. Copy number variations were consistently detected in both NGS and CGH where several chromosomal aneuploidies were correlated to antimony resistance. A major attribute of antimony resistance was a novel terminal deletion of variable length (67kb-204kb) of the polyploid chromosome 31 in the three mutants and was experimentally validated. Terminal deletion in two mutants occurred at the level of inverted repeated sequences in chromosome 31. AQP1 (LmjF.31.0020), a gene encoding for an aquaglyceroporin, which facilitates uptake of trivalent metalloids, was a part of the deleted region. Transfection of AQP1 into resistant mutants rendered them hypersensitive to SbIII. CGH, NGS and Southern blot analysis also highlighted a novel stable, intrachromosomal amplification of a subtelomeric locus on chromosome 34 in one mutant. This region encoded redox enzymes like ascorbate dependent peroxidase (APX) and glucose-6-phosphate dehydrogenase (G6PDH) and overexpression of the genes coding for these enzymes in revertant backgrounds demonstrated resistance to SbIII and protection from reactive oxygen species (ROS) accumulation. Generation of G6PDH null mutant in one revertant exhibited SbIII sensitivity and protection from ROS which were rescued in the add back. Our genomic analyses and parallel functional validation highlighted novel genomic rearrangements, functionally important resistant loci and the implication of new genes in antimony resistance in Leishmania. This submission represents the microarray component of the study 3 biological replicates of each sample
Experiment type
ChIP-chip by array 
Contacts
Angana Mukherjee, Eric Madore, Frederic Raymond
MIAME
PlatformsProtocolsFactorsProcessedRaw
Files
Investigation descriptionE-GEOD-41750.idf.txt
Sample and data relationshipE-GEOD-41750.sdrf.txt
Raw data (1)E-GEOD-41750.raw.1.zip
Processed data (1)E-GEOD-41750.processed.1.zip
Array designA-GEOD-11330.adf.txt
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