Conserved motifs in nuclear genes encoding predicted mitochondrial proteins in Trypanosoma cruzi

Trypanosoma cruzi, the protozoan parasite that causes Chagas’ disease, exhibits peculiar biological features. Among them, the presence of a unique mitochondrion is remarkable. Even though the mitochondrial DNA constitutes up to 25% of total cellular DNA, the structure and functionality of the mitochondrion are dependent on the expression of the nuclear genome. As in other eukaryotes, specific peptide signals have been proposed to drive the mitochondrial localization of a subset of trypanosomatid proteins. However, there are mitochondrial proteins encoded in the nuclear genome that lack of a peptide signal. In other eukaryotes, alternative protein targeting to subcellular organelles via mRNA localization has also been recognized and specific mRNA localization towards the mitochondria has been described. With the aim of seeking for mitochondrial localization signals in T. cruzi, we developed a strategy to build a comprehensive database of nuclear genes encoding predicted mitochondrial proteins (MiNT) in the TriTryps (T. cruzi, T. brucei and L. major). We found that approximately 15% of their nuclear genome encodes mitochondrial products. In T. cruzi the MiNT database reaches 1438 genes and a conserved peptide signal, M(L/F) R (R/S) SS, named TryM-TaPe is found in 60% of these genes, suggesting that the canonical mRNA guidance mechanism is present. In addition, the search for compositional signals in the transcripts of T. cruzi MiNT genes produce a list, being worth to note a conserved nontranslated element represented by the consensus sequence DARRVSG. Taking into account its reported interaction with the T. brucei TRRM3 protein which is enriched in the mitochondrial membrane fraction, we here suggest a putative zip code role for this element. Globally, here we provide an inventory of the mitochondrial proteins in T. cruzi and give evidence for the existence of both peptide and mRNA signals specific to nuclear encoded mitochondrial proteins

Intrinsic DNA curvature in trypanosomes

Background: Trypanosoma cruzi and Trypanosoma brucei are protozoan parasites causing Chagas disease and African sleeping sickness, displaying unique features of cellular and molecular biology. Remarkably, no canonical signals for RNA polymerase II promoters, which drive protein coding genes transcription, have been identified so far. The secondary structure of DNA has long been recognized as a signal in biological processes and more recently, its involvement in transcription initiation in Leishmania was proposed. In order to study whether this feature is conserved in trypanosomatids, we undertook a genome wide search for intrinsic DNA curvature in T. cruzi and T. brucei. Results: Using a region integrated intrinsic curvature (RIIC) scoring that we previously developed, a non-random distribution of sequence-dependent curvature was observed. High RIIC scores were found to be significantly correlated with transcription start sites in T. cruzi, which have been mapped in divergent switch regions, whereas in T. brucei, the high RIIC scores correlated with sites that have been involved not only in RNA polymerase II initiation but also in termination. In addition, we observed regions with high RIIC score presenting in-phase tracts of Adenines, in the subtelomeric regions of the T. brucei chromosomes that harbor the variable surface glycoproteins genes. Conclusions: In both T. cruzi and T. brucei genomes, a link between DNA conformational signals and gene expression was found. High sequence dependent curvature is associated with transcriptional regulation regions. High intrinsic curvature also occurs at the T. brucei chromosome subtelomeric regions where the recombination processes involved in the evasion of the immune host system take place. These findings underscore the relevance of indirect DNA readout in these ancient eukaryotes

Upstream ORFs influence translation efficiency in the parasite Trypanosoma cruzi

It is generally accepted that the presence of ORFs in the 5′ untranslated region of eukaryotic transcripts modulates the production of proteins by controlling the translation initiation rate of the main CDS. In trypanosomatid parasites, which almost exclusively depend on post-transcriptional mechanisms to regulate gene expression, translation has been identified as a key step. However, the mechanisms of control of translation are not fully understood. In the present work, we have annotated the 5′UTRs of the Trypanosoma cruzi genome both in epimastigotes and metacyclic trypomastigotes and, using a stringent classification approach, we identified putative regulatory uORFs in about 9% of the analyzed 5′UTRs. The translation efficiency (TE) and translational levels of transcripts containing putative repressive uORFs were found to be significantly reduced. These findings are supported by the fact that proteomic methods only identify a low number of proteins coded by transcripts containing repressive uORF. We additionally show that AUG is the main translation initiator codon of repressive uORFs in T. cruzi. Interestingly, the decrease in TE is more pronounced when the uORFs overlaps the main CDS. In conclusion, we show that the presence of the uORF and features such as initiation codon and/or location of the uORFs may be acting to fine tune translation levels in these parasites

Draft genome sequence of the UV-Resistant antarctic bacterium Sphingomonas sp. strain UV9

We report the draft genome sequence of the Antarctic UV-resistant bacterium Sphingomonas sp. strain UV9. The strain has a genome size of 4.25 Mb, a 65.62% GC content, and 3,879 protein-coding sequences. Among others, genes encoding the resolving of the DNA damage produced by the UV irradiation were identified

Compositional analysis of flatworm genomes shows strong codon usage biases across all classes

In the present work, we performed a comparative genome-wide analysis of 22 species representative of the main clades and lifestyles of the phylum Platyhelminthes. We selected a set of 700 orthologous genes conserved in all species, measuring changes in GC content, codon, and amino acid usage in orthologous positions. Values of 3rd codon position GC spanned over a wide range, allowing to discriminate two distinctive clusters within freshwater turbellarians, Cestodes and Trematodes respectively. Furthermore, a hierarchical clustering of codon usage data differs remarkably from the phylogenetic tree. Additionally, we detected a synonymous codon usage bias that was more dramatic in extreme GC-poor or GC-rich genomes, i.e., GC-poor Schistosomes preferred to use AT-rich terminated synonymous codons, while GC-rich M. lignano showed the opposite behavior. Interestingly, these biases impacted the amino acidic usage, with preferred amino acids encoded by codons following the GC content trend. These are associated with non-synonymous substitutions at orthologous positions. The detailed analysis of the synonymous and non-synonymous changes provides evidence for a two-hit mechanism where both mutation and selection forces drive the diverse coding strategies of flatworms

Intrinsic DNA curvature in trypanosomes

Trypanosoma cruzi and Trypanosoma brucei are protozoan parasites causing Chagas disease and African sleeping sickness, displaying unique features of cellular and molecular biology. Remarkably, no canonical signals for RNA polymerase II promoters, which drive protein coding genes transcription, have been identified so far. The secondary structure of DNA has long been recognized as a signal in biological processes and more recently, its involvement in transcription initiation in Leishmania was proposed. In order to study whether this feature is conserved in trypanosomatids, we undertook a genome wide search for intrinsic DNA curvature in T. cruzi and T. brucei. Using a region integrated intrinsic curvature (RIIC) scoring that we previously developed, a non-random distribution of sequence-dependent curvature was observed. High RIIC scores were found to be significantly correlated with transcription start sites in T. cruzi, which have been mapped in divergent switch regions, whereas in T. brucei, the high RIIC scores correlated with sites that have been involved not only in RNA polymerase II initiation but also in termination. In addition, we observed regions with high RIIC score presenting in-phase tracts of Adenines, in the subtelomeric regions of the T. brucei chromosomes that harbor the variable surface glycoproteins genes. In both T. cruzi and T. brucei genomes, a link between DNA conformational signals and gene expression was found. High sequence dependent curvature is associated with transcriptional regulation regions. High intrinsic curvature also occurs at the T. brucei chromosome subtelomeric regions where the recombination processes involved in the evasion of the immune host system take place. These findings underscore the relevance of indirect DNA readout in these ancient eukaryotes.https://doi.org/10.1186/s13104-017-2908-

Extensive translational regulation through the proliferative transition of Trypanosoma cruzi revealed by Multi-Omics

Trypanosoma cruzi is the etiological agent for Chagas disease, a neglected parasitic disease in Latin America. Gene transcription control governs the eukaryotic cell replication but is absent in trypanosomatids; thus, it must be replaced by posttranscriptional regulatory events. We investigated the entrance into the T. cruzi replicative cycle using ribosome profiling and proteomics on G1/S epimastigote cultures synchronized with hydroxyurea. We identified 1,784 translationally regulated genes (change > 2, false-discovery rate [FDR]  1.5, FDR < 0.05), respectively. A major translational remodeling accompanied by an extensive proteome change is found, while the transcriptome remains largely unperturbed at the replicative entrance of the cell cycle. The differentially expressed genes comprise specific cell cycle processes, confirming previous findings while revealing candidate cell cycle regulators that undergo previously unnoticed translational regulation. Clusters of genes showing a coordinated regulation at translation and protein abundance share related biological functions such as cytoskeleton organization and mitochondrial metabolism; thus, they may represent posttranscriptional regulons. The translatome and proteome of the coregulated clusters change in both coupled and uncoupled directions, suggesting that complex cross talk between the two processes is required to achieve adequate protein levels of different regulons. This is the first simultaneous assessment of the transcriptome, translatome, and proteome of trypanosomatids, which represent a paradigm for the absence of transcriptional control. The findings suggest that gene expression chronology along the T. cruzi cell cycle is controlled mainly by translatome and proteome changes coordinated using different mechanisms for specific gene groups

Different SNPs in Fasciola hepatica P-glycoprotein from diverse Latin American populations are not associated with Triclabendazole resistance

The use of Triclabendazole for controlling fasciolosis is compromised by increased drug resistance affecting livestock and humans. Although the mode of action of TCBZ is still unknown, putative candidates and markers of resistance have been advanced. A single nucleotide polymorphism (T687 G) in F. hepatica PGP was proposed as marker of resistance in a small scale study of European susceptible and resistant flukes, but the association was not found in Australian samples. The T687 G SNP was absent in more than 40 samples from 2 TCBZ-resistant and 3 susceptible isolates across Latin America here analyzed. While the American samples showed more variable SNPs than the previous ones, none of the SNPs detected showed a marked association with resistance. Analyzing the 42 kb of the FhPGP gene based on RNAseq data highlights that the variation has been underestimated, suggesting that more detailed efforts are needed in order to identify markers of resistance.Fil: Solana, María Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; ArgentinaFil: Domínguez, María Fernanda. Universidad de la República; UruguayFil: Scarcella, Silvana Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; ArgentinaFil: Radio, Santiago. Universidad de la República; UruguayFil: Smircich, Pablo. Universidad de la República; UruguayFil: Fernández, Silvina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; ArgentinaFil: Solana, Hugo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; ArgentinaFil: Tort, José F.. Universidad de la República; Urugua

Survey of transcripts expressed by the invasive juvenile stage of the liver fluke Fasciola hepatica

<p>Abstract</p> <p>Background</p> <p>The common liver fluke <it>Fasciola hepatica </it>is the agent of a zoonosis with significant economic consequences in livestock production worldwide, and increasing relevance to human health in developing countries. Although flukicidal drugs are available, re-infection and emerging resistance are demanding new efficient and inexpensive control strategies. Understanding the molecular mechanisms underlying the host-parasite interaction provide relevant clues in this search, while enlightening the physiological adaptations to parasitism. Genomics and transcriptomics are still in their infancy in <it>F. hepatica</it>, with very scarce information available from the invasive newly excysted juveniles (NEJ). Here we provide an initial glimpse to the transcriptomics of the NEJ, the first stage to interact with the mammalian host.</p> <p>Results</p> <p>We catalogued more than 500 clusters generated from the analysis of <it>F. hepatica </it>juvenile expressed sequence tags (EST), several of them not detected in the adult stage. A set of putative <it>F. hepatica </it>specific transcripts, and a group of sequences conserved exclusively in flatworms were identified. These novel sequences along with a set of parasite transcripts absent in the host genomes are putative new targets for future anti-parasitic drugs or vaccine development.</p> <p>Comparisons of the <it>F. hepatica </it>sequences with other metazoans genomes or EST databases were consistent with the basal positioning of flatworms in the bilaterian phylogeny. Notably, GC content, codon usage and amino acid frequencies are remarkably different in Schistosomes to <it>F. hepatica </it>and other trematodes.</p> <p>Functional annotation of predicted proteins showed a general representation of diverse biological functions. Besides proteases and antioxidant enzymes expected to participate in the early interaction with the host, various proteins involved in gene expression, protein synthesis, cell signaling and mitochondrial enzymes were identified. Differential expression of secreted protease gene family members between juvenile and adult stages may respond to different needs during host colonization.</p> <p>Conclusion</p> <p>The knowledge of the genes expressed by the invasive stage of <it>Fasciola hepatica </it>is a starting point to unravel key aspects of this parasite's biology. The integration of the emerging transcriptomics, and proteomics data and the advent of functional genomics tools in this organism are positioning <it>F. hepatica </it>as an interesting model for trematode biology.</p