Comparative transcriptomic analysis of antimony resistant and susceptible Leishmania infantum lines

Abstract

Background One of the major challenges to leishmaniasis treatment is the emergence of parasites resistant to antimony. To study differentially expressed genes associated with drug resistance, we performed a comparative transcriptomic analysis between wild-type and potassium antimonyl tartrate (SbIII)-resistant Leishmania infantum lines using high-throughput RNA sequencing. Methods All the cDNA libraries were constructed from promastigote forms of each line, sequenced and analyzed using STAR for mapping the reads against the reference genome (L. infantum JPCM5) and DESeq2 for differential expression statistical analyses. All the genes were functionally annotated using sequence similarity search. Results The analytical pipeline considering an adjusted p-value < 0.05 and fold change > 2.0 identified 933 transcripts differentially expressed (DE) between wild-type and SbIII-resistant L. infantum lines. Out of 933 DE transcripts, 504 presented functional annotation and 429 were assigned as hypothetical proteins. A total of 837 transcripts were upregulated and 96 were downregulated in the SbIII-resistant L. infantum line. Using this DE dataset, the proteins were further grouped in functional classes according to the gene ontology database. The functional enrichment analysis for biological processes showed that the upregulated transcripts in the SbIII-resistant line are associated with protein phosphorylation, microtubule-based movement, ubiquitination, host–parasite interaction, cellular process and other categories. The downregulated transcripts in the SbIII-resistant line are assigned in the GO categories: ribonucleoprotein complex, ribosome biogenesis, rRNA processing, nucleosome assembly and translation. Conclusions The transcriptomic profile of L. infantum showed a robust set of genes from different metabolic pathways associated with the antimony resistance phenotype in this parasite. Our results address the complex and multifactorial antimony resistance mechanisms in Leishmania, identifying several candidate genes that may be further evaluated as molecular targets for chemotherapy of leishmaniasis.

Involvement of Leishmania Phosphatases in Parasite Biology and Pathogeny

“…The LiSbR line showed an upregulation of thirty-seven transcripts belonging to the protein phosphorylation category, which includes a DUSP that was 8.93fold upregulated. In addition, a PPP and a PP2C were 2.28-to 17.52-fold upregulated, respectively (Andrade et al, 2020). This is consistent with a previous proteomic analysis that found a major abundance of both enzymes in the Sb III -resistant line (Matrangolo et al, 2013).…”

“…While STPs and PTPs are related to cell signaling (Saxena et al, 2007;Dillon et al, 2015;Alcolea et al, 2016), membrane-bound HAcPs seem to play a role in the response to protein recycling during stress conditions, such as nutrient starvation (Martin et al, 2014;Inbar et al, 2017). Leishmania phosphatases are also upregulated in drug-resistant strains (Matrangolo et al, 2013;Bhandari et al, 2014;Verma et al, 2017;Patino et al, 2019;Andrade et al, 2020). STPs may contribute to parasite resistance by activating ABC transporters, leading to an increase in drug efflux (Bhandari et al, 2014;Verma et al, 2017).…”

“…By using high-throughput RNA sequencing to analyze the transcriptome profiles, significant differences were identified between wild-type and potassium antimonyl tartrate (Sb III )resistant L. infantum lines (abbreviated as LiWTS and LiSbR, respectively) (Andrade et al, 2020). The LiSbR line showed an upregulation of thirty-seven transcripts belonging to the protein phosphorylation category, which includes a DUSP that was 8.93fold upregulated.…”

Perspectives From Systems Biology to Improve Knowledge of Leishmania Drug Resistance

“…Proteomic and genomic analyses of Sb IIIresistant L. infantum mutants identified MRPA as a biomarker and suggested the involvement of chromosome number variations, specific gene amplifications, and SNPs as important features of antimony resistance (Brotherton et al, 2013). The transcriptomic profile showed that many pathways upregulated in L. infantum antimony-resistant lines are associated with protein phosphorylation, microtubule-based movement, protein ubiquitination, stress response, regulation of membrane lipid distribution, proteins involved in RNA metabolism, and other important metabolic pathways (Andrade et al, 2020). Together, these results show that the mechanism of antimony-resistance in Leishmania is complex and multifactorial, identifying several candidate genes that may be further evaluated as molecular targets for chemotherapy of leishmaniasis.…”

“…One possible approach is the integration of public available RNAseq data depicting the resistance phenomena in gene regulatory networks (Andrade et al, 2020). Such approach has demonstrated how genes interact with each other and how changes in their expression levels may result, for example, in different immunological responses promoting distinct disease outcomes including leishmaniasis (Mol et al, 2018).…”