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Splicing is a highly conserved, intricate mechanism intimately linked to transcription elongation, serving as a pivotal regulator of gene expression. Alternative splicing may generate specific transcripts incapable of undergoing translation into proteins, designated as unproductive. A plethora of respiratory viruses, including Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), strategically manipulate the host’s splicing machinery to circumvent antiviral responses. During the infection, SARS-CoV-2 effectively suppresses interferon (IFN) expression, leading to B cell and CD8+ T cell leukopenia, while simultaneously increasing the presence of macrophages and neutrophils in patients with severe COVID-19. In this study, we integrated publicly available omics datasets to systematically analyze transcripts at the isoform level and delineate the nascent-peptide translatome landscapes of SARS-CoV-2-infected human cells. Our findings reveal a hitherto uncharacterized mechanism whereby SARS-CoV-2 infection induces the predominant expression of unproductive splicing isoforms in key IFN signaling genes, interferon-stimulated genes (ISGs), class I MHC genes, and splicing machinery genes, including IRF7, OAS3, HLA-B, and HNRNPH1. In stark contrast, cytokine and chemokine genes, such as IL6, CXCL8, and TNF, predominantly express productive (protein-coding) splicing isoforms in response to SARS-CoV-2 infection. We postulate that SARS-CoV-2 employs a previously unreported tactic of exploiting the host splicing machinery to bolster viral replication and subvert the immune response by selectively upregulating unproductive splicing isoforms from antigen presentation and antiviral response genes. Our study sheds new light on the molecular interplay between SARS-CoV-2 and the host immune system, offering a foundation for the development of novel therapeutic strategies to combat COVID-19.

Yeast communities associated with cacti were studied in three ecosystems of Southeast, Central and North Brazil. A total of 473 yeast strains belonging to 72 species were isolated from 190 samples collected. Cactophilic yeast species were prevalent in necrotic tissues, flowers, fruits and insects of cacti collected in Southeast and North Brazil. Pichia cactophila, Candida sonorensis and species of the Sporopachydermia complex were the most prevalent cactophilic species in Southeast and Central regions. Kodamaea nitidulidarum, Candida restingae and Wickerhamiella cacticola were frequently associated with cactus flowers and fruits. The diversity of yeasts associated with the substrates studied was high. Twenty-one novel species were found. One is described here as Kluyveromyces starmeri sp. nov. based on 21 isolates obtained from necrotic tissues, flowers, fruits and associated insects of the columnar cacti Cereus saddianus, Micranthocereus dolichospermaticus and Pilosocereus arrabidae in two different ecosystems in Brazil. Phylogenetic analyses of sequences encoding the gene of the small subunit (SSU) rRNA gene, the internal transcribed spacer, the 5.8S rRNA gene and the D1/D2 domains of the large subunit (LSU) rRNA showed that the species is related to Kluyveromyces dobzhanskii, Kluyveromyces lactis and Kluyveromyces marxianus. Phylogenomic analyses based on 1264 conserved genes shared among the new species and 19 other members of the Saccharomycetaceae confirmed this phylogenetic relationship. The holotype is K. starmeri sp. nov. CBS 16103T (=UFMG-CM-Y3682T ). The Mycobank number is MB 836817.

The snow-covered surfaces of Antarctica comprise an extreme environment that favors the development of life forms with adaptations to adverse low-temperature habitats. The ability to survive and such temperatures might involve the production of antifreeze proteins and ice-binding proteins that attenuate the effects of intense cold temperatures. He, we sequenced and reconstructed the nuclear and mitochondrial genomes of the endemic Antarctic fungus Antarctomyces pellizariae UFMGCB 12416. We then have identified a putative ice-binding protein-coding gene, mapped the presence of secondary metabolite gene clusters, and reconstructed the phylogenetic relationships of a A. pellizariae with others Leotiomycetes from the alignment of hundreds of orthologous single-copy proteins. Our results will deepen the understanding of microbial ice-binding proteins and the genomic aspects of psychrophilic fungi. Dataset The GenBank/EMBL/DDBJ accession number for the gene sequence of ice-binding protein from A. pellizariae determined in this study is MN867686. The Whole Genome Shotgun project of strain A. pellizariae UFMGCB 12416 has been deposited at DDBJ/ENA/GenBank under accession WCAA00000000. The version described in this paper is version WCAA01000000. The mitochondrial genome has been deposited under accession MT197497.

Aim: GDF15 levels are a biomarker for metformin use. We performed the functional annotation of noncoding genome-wide association study (GWAS) SNPs for GDF15 levels and the Genotype-Tissue Expression (GTEx)-expression quantitative trait loci (eQTLs) for GDF15 expression within metformin-activated enhancers around GDF15. Materials & methods: These enhancers were identified using chromatin immunoprecipitation followed by sequencing data for active (H3K27ac) and silenced (H3K27me3) histone marks on human hepatocytes treated with metformin, Encyclopedia of DNA Elements data and cis-regulatory elements assignment tools. Results: The GWAS lead SNP rs888663, the SNP rs62122429 associated with GDF15 levels in the Outcome Reduction with Initial Glargine Intervention trial, and the GTEx-expression quantitative trait locus rs4808791 for GDF15 expression in whole blood are located in a metformin-activated enhancer upstream of GDF15 and tightly linked in Europeans and East Asians. Conclusion: Noncoding variation within a metformin-activated enhancer may increase GDF15 expression and help to predict GDF15 levels.

Trypanosoma cruzi, the causative agent of Chagas disease, has a complex life cycle that requires the adaptation to different environments. In the absence of traditional mechanisms for regulation of gene expression, this parasite relies on posttranscriptional control events, which allow the progression of its life cycle in different hosts and stress conditions. In this context, different stress conditions trigger the aggregation of RNA-binding proteins and their target mRNAs into cytoplasmic foci known as RNA granules, which act as RNA-sorting centers. In this study, we have characterized the T. cruzi RNA-binding protein ALBA30 during nutritional stress conditions. Using a recombinant form of TcALBA30 to facilitate its detection (rTcALBA30), we showed that this protein resides in the cytoplasm in normal growth conditions but is recruited into cytoplasmic foci after starvation. Moreover, evaluation of rTcALBA30 in parasites that reached the stationary phase of growth also showed the recruitment of this protein into cytoplasmic foci. Our results indicate that, similar to TbALBA3, TcALBA30 aggregates into stress granules in parasites submitted to nutritional stress.

Acute rejection (AR) is a process triggered via the recognition of grafted organ-derived antigens by the immune system, which could present as a life-threatening condition. In the context of a kidney transplant, despite improvement with immunosuppressive therapies, AR maintains a significant incidence of 10%, and currently available drugs generally act in similar and canonical pathways of lymphocyte activation. This prompted the research for different approaches to identify potential novel targets that could improve therapeutic interventions. Here, we conducted a transcriptome analysis comparing groups of acute rejection (including T cell-mediated rejection and antibody-mediated rejection) to stable grafts that included differentially expressed genes, transcription factor and kinase enrichment, and Gene Set Enrichment Analysis. These analyses revealed inflammasome enhancement in rejected grafts and AIM2 as a potential component linked to acute rejection, presenting a positive correlation to T-cell activation and a negative correlation to oxidative phosphorylation metabolism. Also, the AIM2 expression showed a global accuracy in discerning acute rejection grafts (area under the curve (AUC) = 0.755 and 0.894, p < 0.0001), and meta-analysis comprising different studies indicated a considerable enhancement of AIM2 in rejection (standardized mean difference (SMD) = 1.45, [CI 95%, 1.18 to 1.71]), especially for T cell-mediated rejection (TCMR) (SMD = 2.01, [CI 95%, 1.58 to 2.45]). These findings could guide future studies of AIM2 as either an adjuvant target for immunosuppression or a potential biomarker for acute rejection and graft survival.

Summary Thousands of biomedical scientific articles, including those describing genes associated with human diseases, are published every week. Computational methods such as text mining and machine learning algorithms are now able to automatically detect these associations. In this study, we used a cognitive computing text-mining application to construct a knowledge network comprising 3,723 genes and 99 diseases. We then tracked the yearly changes on these networks to analyze how our knowledge has evolved in the past 30 years. Our systems approach helped to unravel the molecular bases of diseases and detect shared mechanisms between clinically distinct diseases. It also revealed that multi-purpose therapeutic drugs target genes that are commonly associated with several psychiatric, inflammatory, or infectious disorders. By navigating this knowledge tsunami, we were able to extract relevant biological information and insights about human diseases., Graphical abstract, Highlights • Over 3,700 genes were associated with 99 human diseases in the scientific literature • The knowledge on human disease genes increased exponentially in the past 30 years • Knowledge networks revealed genes shared by very different diseases • Hub genes are known drug targets with drug repositioning potential, Molecular network; Bioinformatics; Association analysis; Systems biology

Background PTEN is the most frequently lost tumor suppressor in primary prostate cancer (PCa) and its loss is associated with aggressive disease. However, the transcriptional changes associated with PTEN loss in PCa have not been described in detail. In this study, we highlight the transcriptional changes associated with PTEN loss in PCa. Methods Using a meta-analysis approach, we leveraged two large PCa cohorts with experimentally validated PTEN and ERG status by Immunohistochemistry (IHC), to derive a transcriptomic signature of PTEN loss, while also accounting for potential confounders due to ERG rearrangements. This signature was expanded to lncRNAs using the TCGA quantifications from the FC-R2 expression atlas. Results The signatures indicate a strong activation of both innate and adaptive immune systems upon PTEN loss, as well as an expected activation of cell-cycle genes. Moreover, we made use of our recently developed FC-R2 expression atlas to expand this signature to include many non-coding RNAs recently annotated by the FANTOM consortium. Highlighting potential novel lncRNAs associated with PTEN loss and PCa progression. Conclusion We created a PCa specific signature of the transcriptional landscape of PTEN loss that comprises both the coding and an extensive non-coding counterpart, highlighting potential new players in PCa progression. We also show that contrary to what is observed in other cancers, PTEN loss in PCa leads to increased activation of the immune system. These findings can help the development of new biomarkers and help guide therapy choices.

DNA topoisomerases are enzymes that modulate DNA topology. Among them, topoisomerase 3α is engaged in genomic maintenance acting in DNA replication termination, sister chromatid separation, and dissolution of recombination intermediates. To evaluate the role of this enzyme in Trypanosoma cruzi, the etiologic agent of Chagas disease, a topoisomerase 3α knockout parasite (TcTopo3α KO) was generated, and the parasite growth, as well as its response to several DNA damage agents, were evaluated. There was no growth alteration caused by the TcTopo3α knockout in epimastigote forms, but a higher dormancy rate was observed. TcTopo3α KO trypomastigote forms displayed reduced invasion rates in LLC-MK2 cells when compared with the wild-type lineage. Amastigote proliferation was also compromised in the TcTopo3α KO, and a higher number of dormant cells was observed. Additionally, TcTopo3α KO epimastigotes were not able to recover cell growth after gamma radiation exposure, suggesting the involvement of topoisomerase 3α in homologous recombination. These parasites were also sensitive to drugs that generate replication stress, such as cisplatin (Cis), hydroxyurea (HU), and methyl methanesulfonate (MMS). In response to HU and Cis treatments, TcTopo3α KO parasites showed a slower cell growth and was not able to efficiently repair the DNA damage induced by these genotoxic agents. The cell growth phenotype observed after MMS treatment was similar to that observed after gamma radiation, although there were fewer dormant cells after MMS exposure. TcTopo3α KO parasites showed a population with sub-G1 DNA content and strong γH2A signal 48 h after MMS treatment. So, it is possible that DNA-damaged cell proliferation due to the absence of TcTopo3α leads to cell death. Whole genome sequencing of MMS-treated parasites showed a significant reduction in the content of the multigene families DFG-1 and RHS, and also a possible erosion of the sub-telomeric region from chromosome 22, relative to non-treated knockout parasites. Southern blot experiments suggest telomere shortening, which could indicate genomic instability in TcTopo3α KO cells owing to MMS treatment. Thus, topoisomerase 3α is important for homologous recombination repair and replication stress in T. cruzi, even though all the pathways in which this enzyme participates during the replication stress response remains elusive.

The Pseudomonas putida group comprises strains with biotechnological and clinical relevance. P. alloputida was proposed as a new species and highlighted the misclassification of P. putida. Nevertheless, the population structure of P. alloputida remained unexplored. We retrieved 11,025 Pseudomonas genomes and used P. alloputida Kh7T to delineate the species. The P. alloputida population structure comprises at least 7 clonal complexes (CCs). Clinical isolates are mainly found in CC4 and acquired resistance genes are present at low frequency in plasmids. Virulence profiles support the potential of CC7 members to outcompete other plant or human pathogens through a type VI secretion system. Finally, we found that horizontal gene transfer had an important role in shaping the ability of P. alloputida to bioremediate aromatic compounds such as toluene. Our results provide the grounds to understand P. alloputida genetic diversity and its potential for biotechnological applications.

RNA modifications are dynamic chemical entities that expand the RNA lexicon and regulate RNA fate. The most abundant modification present in mRNAs, N6-methyladenosine (m6A), has been implicated in neurogenesis and memory formation. However, whether additional RNA modifications may be playing a role in neuronal functions and in response to environmental queues is largely unknown. Here we characterize the biochemical function and cellular dynamics of two human RNA methyltransferases previously associated with neurological dysfunction, TRMT1 and its homolog, TRMT1-like (TRMT1L). Using a combination of next-generation sequencing, LC-MS/MS, patient-derived cell lines and knockout mouse models, we confirm the previously reported dimethylguanosine (m2,2G) activity of TRMT1 in tRNAs, as well as reveal that TRMT1L, whose activity was unknown, is responsible for methylating a subset of cytosolic tRNAAla(AGC) isodecoders at position 26. Using a cellular in vitro model that mimics neuronal activation and long term potentiation, we find that both TRMT1 and TRMT1L change their subcellular localization upon neuronal activation. Specifically, we observe a major subcellular relocalization from mitochondria and other cytoplasmic domains (TRMT1) and nucleoli (TRMT1L) to different small punctate compartments in the nucleus, which are as yet uncharacterized. This phenomenon does not occur upon heat shock, suggesting that the relocalization of TRMT1 and TRMT1L is not a general reaction to stress, but rather a specific response to neuronal activation. Our results suggest that subcellular relocalization of RNA modification enzymes may play a role in neuronal plasticity and transmission of information, presumably by addressing new targets. NJ was supported by a UNSW International PhD fellowship. SC is supported by a fellowship from ”la Caixa'' Foundation (LCF/BQ/DI19/11730036). This work was supported by NHMRC funds (Project Grant APP1070631 to JSM), funds from the Australian Research Council (DP180103571 to EMN) and funds from the Garvan Young Investigator Award (to NS). This work was partly supported by the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) (PGC2018-098152-A-100 to EMN).The mass spectrometric analyses shown in Figure S3 were performed in the CRG/UPF Proteomics Unit which is part of the of Proteored, PRB3 and is supported by grant PT17/0019, of the PE I+D+i 2013-2016, funded by ISCIII and ERDF

Members of the Pseudomonas genus inhabit a wide variety of environments, which is correlated with their versatile genome and metabolic capacity. Pseudomonas putida is a saprophytic soil bacterium, known to bioremediate toxic compounds and to promote plant growth. Nevertheless, some clinical P. putida strains have been reported, raising concerns about the safety of this species for biotechnology applications. Despite the ability to occupy different niches, the population structure of P. putida remains unclear. Here, we report the population structure of P. putida for the first time, which is essential to better investigate its biology. We retrieved 11,025 Pseudomonas genomes and used P. putida KT2440 as a type strain to define P. putida sensu stricto, as well as to estimate its population structure and pangenome. By using network and average nucleotide identity analyses, we found that 90 of the 138 deposited P. putida genomes were misclassified, including some reference genomes. Further, we found 23 additional P. putida genomes that were not designated as such. P. putida has an open pangenome dominated by low-frequency genes. The population structure of P. putida is composed by at least 7 clonal complexes (CCs). Clinical isolates are not restricted to a specific CC, although CC4 has 6 out the 9 clinical isolates. Acquired resistance genes are present at low frequency, particularly in plasmids from clinical isolates. Virulence profiles support the higher potential of CC7 members to outcompete other plant or human pathogens through a type VI secretion system. P. putida lacks key genes for indole-3-acetic acid production, indicating an incomplete or non-functional pathway. Finally, we found that horizontal gene transfer had an important role in shaping the ability of P. putida to bioremediate aromatic compounds such as toluene. Our results provide the grounds to understand P. putida genetic diversity, which is key to assess its safety for environmental and biotechnology applications.

RNA modifications are dynamic chemical entities that regulate RNA fate, and an avenue for environmental response in neuronal function. However, which RNA modifications may be playing a role in neuronal plasticity and environmental responses is largely unknown. Here we characterize the biochemical function and cellular dynamics of two human RNA methyltransferases previously associated with neurological dysfunction, TRMT1 and its homolog, TRMT1-like (TRMT1L). Using a combination of next-generation sequencing, LC-MS/MS, patient-derived cell lines and knockout mouse models, we confirm the previously reported dimethylguanosine (m 2,2 G) activity of TRMT1 in tRNAs, as well as reveal that TRMT1L, whose activity was unknown, is responsible for methylating a subset of cytosolic tRNA Ala (AGC) isoacceptors at position 26. Using a cellular in vitro model that mimics neuronal activation and long term potentiation, we find that both TRMT1 and TRMT1L change their subcellular localization upon neuronal activation. Specifically, we observe a major subcellular relocalization from mitochondria and other cytoplasmic domains (TRMT1) and nucleoli (TRMT1L) to different small punctate compartments in the nucleus, which are as yet uncharacterized. This phenomenon does not occur upon heat shock, suggesting that the relocalization of TRMT1 and TRMT1L is not a general reaction to stress, but rather a specific response to neuronal activation. Our results suggest that subcellular relocalization of RNA modification enzymes play a role in neuronal plasticity and transmission of information, presumably by addressing new targets.

PTEN is the most frequently lost tumor suppressor in primary prostate cancer (PCa) and its loss is associated with aggressive disease. However, the transcriptional changes associated with PTEN loss in PCa have not been described in detail. Here, we applied a meta-analysis approach, leveraging two large PCa cohorts with experimentally validated PTEN and ERG status, to derive a transcriptomic signature ofPTENloss, while also accounting for potential confounders due toERGrearrangements. Strikingly, the signature indicates a strong activation of both innate and adaptive immune systems uponPTENloss, as well as an expected activation of cell-cycle genes. Moreover, we made use of our recently developed FC-R2 expression atlas to expand this signature to include many non-coding RNAs recently annotated by the FANTOM consortium. With this resource, we analyzed the TCGA-PRAD cohort, creating a comprehensive transcriptomic landscape ofPTENloss in PCa that comprises both the coding and an extensive non-coding counterpart.

The protozoanTrypanosoma cruzi(T. cruzi) is a well-adapted parasite to mammalian hosts and the pathogen of Chagas disease in humans. As both host andT. cruziare highly genetically diverse, many variables come into play during infection, making disease outcomes difficult to predict. One important challenge in the field of Chagas disease research is determining the main factors leading to parasite establishment in the chronic stage in some organs, mainly the heart and/or digestive system. Our group previously showed that distinct strains ofT. cruzi(JG and Col1.7G2) acquired differential tissue distribution in the chronic stage in dually-infected BALB/c mice. To investigate changes in the host triggered by the two distinctT. cruzistrains, we assessed the gene expression profile of BALB/c mouse hearts infected with either JG, Col1.7G2 or an equivalent mixture of both parasites during the initial phase of infection. This study demonstrates a clear distinction in host gene expression modulation by both parasites. Col1.7G2 strongly activated Th1-polarized immune signature genes, whereas JG showed only minor activation of the host immune response. Moreover, JG strongly reduced the expression of genes for ribosomal proteins and mitochondrial proteins related to the electron transport chain. Interestingly, evaluation of gene expression in mice inoculated with the mixture of parasites showed expression profiles for both up- and down-regulated genes, indicating the coexistence of both parasite strains in the heart during the acute phase. This study suggests that different strains ofT. cruzimay be distinguished by their efficiency in activating the immune system, modulating host energy and reactive oxygen species production and decreasing protein synthesis during early infection, which may be crucial in defining parasite persistence in specific organs.Author SummaryThe causative agent of Chagas disease,Trypanosoma cruzi, retains high genetic diversity, and its populations vary greatly across geographic locations. TheT. cruzimammalian hosts, including humans, also have high genetic variation, making it difficult to predict the disease outcome. Accordingly, this variability must be taken into account in several studies aiming to interrogate the effect of polyparasitism in drug trials, vaccines, diagnosis or basic research. Therefore, there is a growing need to consider the interaction between the pathogen and the host immune system in mixed infections. In the present work, we present an in-depth analysis of the gene expression of hearts from BALB/c mice infected with Col1.7G2 and JG alone or a mixture of both strains. Col1.7G2 induced a higher Th1 inflammatory response, while JG exhibited a weaker activation of immune response genes. Furthermore, JG-infected mice showed a notable reduction in the expression of genes responsible for mitochondrial oxidative phosphorylation and protein synthesis. Interestingly, the mixture-infected group displayed changes in gene expression as caused by both strains. Overall, we provided new insights into the host-pathogen interaction in the context of single and dual infection, showing remarkable differences in host gene expression modulation by twoT. cruzistrains.

Long noncoding RNAs (lncRNAs) have emerged as key coordinators of biological and cellular processes. Characterizing lncRNA expression across cells and tissues is key to understanding their role in determining phenotypes, including human diseases. We present here FC-R2, a comprehensive expression atlas across a broadly defined human transcriptome, inclusive of over 109,000 coding and noncoding genes, as described in the FANTOM CAGE-Associated Transcriptome (FANTOM-CAT) study. This atlas greatly extends the gene annotation used in the originalrecount2resource. We demonstrate the utility of the FC-R2 atlas by reproducing key findings from published large studies and by generating new results across normal and diseased human samples. In particular, we (a) identify tissue-specific transcription profiles for distinct classes of coding and noncoding genes, (b) perform differential expression analysis across thirteen cancer types, identifying novel noncoding genes potentially involved in tumor pathogenesis and progression, and (c) confirm the prognostic value for several enhancer lncRNAs expression in cancer. Our resource is instrumental for the systematic molecular characterization of lncRNA by the FANTOM6 Consortium. In conclusion, comprised of over 70,000 samples, the FC-R2 atlas will empower other researchers to investigate functions and biological roles of both known coding genes and novel lncRNAs.

Psychiatric and neurological disorders (PNDs) affect millions worldwide and only a few drugs achieve complete therapeutic success in the treatment of these disorders. Due to the high cost of developing novel drugs, drug repositioning represents a promising alternative method of treatment. In this manuscript, we used a network medicine approach to investigate the molecular characteristics of PNDs and identify novel drug candidates for repositioning. Using IBM Watson for Drug Discovery, a powerful machine learning text-mining application, we built knowledge networks containing connections between PNDs and genes or drugs mentioned in the scientific literature published in the past 50 years. This approach revealed several drugs that target key PND-related genes, which have never been used to treat these disorders to date. We validate our framework by detecting drugs that have been undergoing clinical trial for treating some of the PNDs, but have no published results in their support. Our data provides comprehensive insights into the molecular pathology of PNDs and offers promising drug repositioning candidates for follow-up trials.

Two isolates representing a new species of Scheffersomyces were isolated from rotting wood samples collected in an Amazonian forest ecosystem in Brazil. Analysis of the sequences of the D1/D2 domains showed that this new species is phylogenetically related to Scheffersomyces NYMU 15730, a species without a formal description, and the two are in an early emerging position with respect to the xylose-fermenting subclade containing Scheffersomyces titanus and Scheffersomyces stipitis. Phylogenomic analyses using 474 orthologous genes placed the new species in an intermediary position between Scheffersomyces species and the larger genus Spathaspora and the Candida albicans/Lodderomyces clade. The novel species, Scheffersomyces stambukii f.a., sp. nov., is proposed to accommodate these isolates. The type strain of Scheffersomyces stambukii sp. nov. is UFMG-CM-Y427T (=CBS 14217T). The MycoBank number is MB 824093. In addition, we studied the xylose metabolism of this new species.

Trypanosoma cruzi is the etiological agent of Chagas disease, a public health challenge due to its morbidity and mortality rates, which affects around 6-7 million people worldwide. Symptoms, response to chemotherapy, and the course of Chagas disease are greatly influenced by T. cruzi‘s intra-specific variability. Thus, DNA mutations in this parasite possibly play a key role in the wide range of clinical manifestations and in drug sensitivity. Indeed, the environmental conditions of oxidative stress faced by T. cruzi during its life cycle can generate genetic mutations. However, the lack of an established experimental design to assess mutation rates in T. cruzi precludes the study of conditions and mechanisms that potentially produce genomic variability in this parasite. We developed an assay that employs a reporter gene that, once mutated in specific positions, convert G418-sensitive into G418-insenstitive T. cruzi. We were able to determine the frequency of DNA mutations in T. cruzi exposed and non-exposed to oxidative insults assessing the number of colony-forming units in solid selective media after plating a defined number of cells. We verified that T. cruzi‘s spontaneous mutation frequency was comparable to those found in other eukaryotes, and that exposure to hydrogen peroxide promoted a two-fold increase in T. cruzi‘s mutation frequency. We hypothesize that genetic mutations in T. cruzi can arise from oxidative insults faced by this parasite during its life cycle.

Two yeast isolates producing asci-containing elongate ascospores with curved ends typical of the genus Spathaspora were isolated from rotting wood samples collected in an Atlantic rainforest ecosystem in Brazil. Phylogenetic analysis of the LSU rRNA gene D1/D2 domain sequences demonstrated that the strains represent a new species and placed it next to Candida blackwellae, in a clade that also contains Candida albicans and Candida dubliniensis. Other sequences of the ribosomal gene cluster supported same placementin the same clade, and a phylogenomic analysis placed this new species in an early emerging position relative to the larger C. albicans/Lodderomyces clade. One interpretation is that the genus Spathaspora is, in fact, paraphyletic. In conformity with this view, we propose the novel species Spathaspora boniae sp. nov. to accommodate the isolates. The type strain of Spathaspora boniae sp. nov. is UFMG-CM-Y306T (=CBS 13262T). The MycoBank number is MB 821297. A detailed analysis of xylose metabolism was conducted for the new species.

SUMMARYTrypanosoma cruziis exposed to oxidative stresses during its life cycle, and amongst the strategies employed by this parasite to deal with these situations sits a peculiar trypanothione-dependent antioxidant system. Remarkably,T. cruzi’s antioxidant repertoire does not include catalase. In an attempt to shed light on what are the reasons by which this parasite lacks this enzyme, aT. cruzicell line stably expressing catalase showed an increased resistance to hydrogen peroxide (H2O2) when compared with wild-type cells. Interestingly, preconditioning carried out with low concentrations of H2O2led untransfected parasites to be as much resistant to this oxidant as cells expressing catalase, but did not induce the same level of increased resistance in the latter ones. Also, presence of catalase decreased trypanothione reductase and increased superoxide dismutase levels inT. cruzi, resulting in higher levels of residual H2O2after challenge with this oxidant. Although expression of catalase contributed to elevated proliferation rates ofT. cruziinRhodnius prolixus, it failed to induce a significant increase of parasite virulence in mice. Altogether, these results indicate that the absence of a gene encoding catalase inT. cruzihas played an important role in allowing this parasite to develop a shrill capacity to sense and overcome oxidative stress.

Long non-coding RNAs (lncRNAs) have emerged as key coordinators of biological and cellular processes. Characterizing lncRNA expression across cells and tissues is key to understanding their role in determining phenotypes including human diseases. We present here FC-R2, a comprehensive expression atlas across a broadly-defined human transcriptome, inclusive of over 109,000 coding and non-coding genes, as described in the FANTOM CAGE-Associated Transcriptome (FANTOM-CAT) study. This atlas greatly extends the gene annotation used in the original recount2 resource. We demonstrate the utility of the FC-R2 atlas by reproducing key findings from published large studies and by generating new results across normal and diseased human samples. In particular, we (a) identify tissue specific transcription profiles for distinct classes of coding and non-coding genes, (b) perform differential expression analyses across thirteen cancer types, providing new insights linking promoter and enhancer lncRNAs expression to tumor pathogenesis, and (c) confirm the prognostic value of several enhancers in cancer. Comprised of over 70,000 samples, the FC-R2 atlas will empower other researchers to investigate functions and biological roles of both known coding genes and novel lncRNAs. Most importantly, access to the FC-R2 atlas is available from https://jhubiostatistics.shinyapps.io/recount/, the recount Bioconductor package, and http://marchionnilab.org/fcr2.html.

Antarctica is a reservoir of microbial biodiversity that has only partially been characterized. The same extreme conditions that hinder the scientific investigation of the continent have also selected organisms with remarkable adaptations and promising biotechnological applications. Fungi are widely recognized as being capable of producing a variety of compounds, and though many have already been isolated and characterized from diverse Antarctic environments, a deeper understanding of these organisms and the molecular mechanisms responsible for their capabilities is needed. The recent advances in sequencing and bioinformatics analysis tools are revolutionizing the life sciences and are already being applied to Antarctic bioprospection. Despite being regarded as a separate field, in which most researchers would rather not venture, bioinformatics presents a new frontier, especially to microbiologists. Comprehension of bioinformatics tools and techniques has much to add to any microbiological analysis. This chapter intends to familiarize the reader with the concepts and logic behind diverse bioinformatics methodologies and to present the omics studies performed so far on Antarctic fungal biodiversity, with a focus on the sequencing, assembly, and investigation of genomes.

Made available in DSpace on 2018-12-11T16:40:58Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-03-15 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Phospholipases A2 are major components of snake venoms (svPLA2s) and are able to induce multiple local and systemic deleterious effects upon envenomation. Several snake species are provided with svPLA2 inhibitors (sbPLIs) in their circulating blood, which confer a natural resistance against the toxic components of homologous and heterologous venoms. The sbPLIs belong to any of three structural classes named α, β and γ. In the present study, we identified, characterized and performed structural and evolutionary analyses of sbαPLIs transcripts and the encoded proteins, in the most common Latin American pit vipers belonging to Crotalus, Bothrops and Lachesis genera. Mutation data indicated that sbαPLIs from Latin American snakes might have evolved in an accelerated manner, similarly to that reported for sbαPLIs from Asian snakes, and possibly co-evoluted with svPLA2s in response to the diversity of target enzymes. The importance of sbαPLI trimerization for the effective binding and inhibition of acidic svPLA2s is discussed and conserved cationic residues located at the central pore of the inhibitor trimer are suggested to be a significant part of the binding site of sbαPLIs to acidic svPLA2s. Our data contribute to the current body of knowledge on the structural and evolutionary characteristics of sbPLIs, in general, and may assist in the future development of selective inhibitors for secretory PLA2 from several sources. Fundação Ezequiel Dias (FUNED) Departamento de Bioquímica e Imunologia Universidade Federal de Minas Gerais (UFMG) Departamento de Física e Biofísica Instituto de Biociências Universidade Estadual Paulista (UNESP) Departamento de Física e Biofísica Instituto de Biociências Universidade Estadual Paulista (UNESP) CNPq: 1810/11

Detection of genetic exchange has been a limiting factor to deepen the knowledge on the mechanisms by which Trypanosoma cruzi is able to generate progeny and genetic diversity. Here we show that incorporation of halogenated thymidine analogues, followed by immunostaining, is a reliable method not only to detect T. cruzi fused-cell hybrids, but also to quantify their percentage in populations of this parasite. Through this approach, we were able to detect and quantify fused-cell hybrids of T. cruzi clones CL Brener and Y. Given the increased detection of fused-cell hybrids in naturally-occurring hybrid CL Brener strain, which displays increased levels of RAD51 and BRCA2 transcripts, we further investigated the role of Rad51 – a recombinase involved in homologous recombination – in the process of genetic exchange. We also verified that the detection of fused-cell hybrids in T. cruzi overexpressing RAD51 is increased when compared to wild-type cells, suggesting a key role for Rad51 either in the formation or in the stabilization of fused-cell hybrids in this organism.

Background: The hemibiotrophic pathogens Moniliophthora perniciosa (witches' broom disease) and Moniliophthora roreri (frosty pod rot disease) are among the most important pathogens of cacao. Moniliophthora perniciosa has a broad host range and infects a variety of meristematic tissues in cacao plants, whereas M. roreri infects only pods of Theobroma and Herrania genera. Comparative pathogenomics of these fungi is essential to understand Moniliophthora infection strategies, therefore the detection and in silico functional characterization of effector candidates are important steps to gain insight on their pathogenicity. Results: Candidate secreted effector proteins repertoire were predicted using the genomes of five representative isolates of M. perniciosa subpopulations (three from cacao and two from solanaceous hosts), and one representative isolate of M. roreri from Peru. Many putative effectors candidates were identified in M. perniciosa: 157 and 134 in cacao isolates from Bahia, Brazil; 109 in cacao isolate from Ecuador, 92 and 80 in wild solanaceous isolates from Minas Gerais (Lobeira) and Bahia (Caiçara), Brazil; respectively. Moniliophthora roreri showed the highest number of effector candidates, a total of 243. A set of eight core effectors were shared among all Moniliophthora isolates, while others were shared either between the wild solanaceous isolates or among cacao isolates. Mostly, candidate effectors of M. perniciosa were shared among the isolates, whereas in M. roreri nearly 50% were exclusive to the specie. In addition, a large number of cell wall-degrading enzymes characteristic of hemibiotrophic fungi were found. From these, we highlighted the proteins involved in cell wall modification, an enzymatic arsenal that allows the plant pathogens to inhabit environments with oxidative stress, which promotes degradation of plant compounds and facilitates infection. Conclusions: The present work reports six genomes and provides a database of the putative effectorome of Moniliophthora, a first step towards the understanding of the functional basis of fungal pathogenicity. © 2018 The Author(s). This work was done in the frame of the International Consortium in Advanced Biology (CIBA; https://www.ciba-network.org). The authors thank the Molecular Plant Pathology Laboratory and the Plant Pathology Laboratory at INIAP personnel for their assistance in obtaining the DNAs, Dr Carmen Suarez Capello for her kind assistance in Ecuador, and the Núcleo de Biologia Computacional e Gestão de Informações Biotecnológicas - UESC (NBCGIB), and Copenhague University for providing bioinformatics facility. Data sets were processed in sagarana HPC cluster, CPAD-ICB-UFMG. The authors would also like to thank Dr. Claudia Fortes Ferreira (Embrapa CNPMF, Brazil) and Dr. Raul Renné Valle (CEPLAC/CEPEC, Brazil) for English language revision. We are also grateful to Ivanna Michelle Meraz Pérez for helping translating an early version of this manuscript and to the anonymous reviewers who provided helpful comments to our work. KPG, FM and CPP were supported by research fellowship Pq-1 from CNPq. National Council for Scientific Development (CNPq) n° 311759/2014–9. CSB acknowledges FAPESB (Foundation for Research Support of the State of Bahia) for supporting her with a research assistantship during her Master’s Programme.

Spliced leader dependent trans-splicing (SLTS) has been described as an important RNA regulatory process that occurs in different organisms, including the trematode Schistosoma mansoni. We identified more than seven thousand putative SLTS sites in the parasite, comprising genes with a wide spectrum of functional classes, which underlines the SLTS as a ubiquitous mechanism in the parasite. Also, SLTS gene expression levels span several orders of magnitude, showing that SLTS frequency is not determined by the expression level of the target gene, but by the presence of particular gene features facilitating or hindering the trans-splicing mechanism. Our in-depth investigation of SLTS events demonstrates widespread alternative trans-splicing (ATS) acceptor sites occurring in different regions along the entire gene body, highlighting another important role of SLTS generating alternative RNA isoforms in the parasite, besides the polycistron resolution. Particularly for introns where SLTS directly competes for the same acceptor substrate with cis-splicing, we identified for the first time additional and important features that might determine the type of splicing. Our study substantially extends the current knowledge of RNA processing by SLTS in S. mansoni, and provide basis for future studies on the trans-splicing mechanism in other eukaryotes.

In Trypanosoma cruzi, the etiologic agent of Chagas disease, Rad51 (TcRad51) is a central enzyme for homologous recombination. Here we describe the different roles of TcRad51 in DNA repair. Epimastigotes of T. cruzi overexpressing TcRAD51 presented abundant TcRad51-labeled foci before gamma irradiation treatment, and a faster growth recovery when compared to single-knockout epimastigotes for RAD51. Overexpression of RAD51 also promoted increased resistance against hydrogen peroxide treatment, while the single-knockout epimastigotes for RAD51 exhibited increased sensitivity to this oxidant agent, which indicates a role for this gene in the repair of DNA oxidative lesions. In contrast, TcRad51 was not involved in the repair of crosslink lesions promoted by UV light and cisplatin treatment. Also, RAD51 single-knockout epimastigotes showed a similar growth rate to that exhibited by wild-type ones after treatment with hydroxyurea, but an increased sensitivity to methyl methane sulfonate. Besides its role in epimastigotes, TcRad51 is also important during mammalian infection, as shown by increased detection of T. cruzi cells overexpressing RAD51, and decreased detection of single-knockout cells for RAD51, in both fibroblasts and macrophages infected with amastigotes. Besides that, RAD51-overexpressing parasites infecting mice also presented increased infectivity and higher resistance against benznidazole. We thus show that TcRad51 is involved in the repair of DNA double strands breaks and oxidative lesions in two different T. cruzi developmental stages, possibly playing an important role in the infectivity of this parasite., Author summary Trypanosoma cruzi is the causative agent of Chagas disease, a tropical neglected illness that affects 6 million people worldwide, mostly in Latin America. Our research group focuses on the elucidation of DNA repair and metabolism of T. cruzi, and on the possible implications of those mechanisms in both parasite genetic diversity and Chagas disease development. In this work, we investigated the involvement of homologous recombination in the oxidation-induced damage response, and in DNA repair, in T. cruzi cells after gamma radiation exposure. We also examined whether TcRad51 –a key protein for homologous recombination–could be an important factor for T. cruzi’s infectivity. We generated genetically-modified T. cruzi cells for RAD51 and studied the in vitro and in vivo infectivity of these mutant cells. Our results indicate that TcRad51 is a key protein involved in the repair of oxidative and DNA double-strand breaks lesions in two crucial developmental forms of T. cruzi.

Made available in DSpace on 2018-11-26T15:44:34Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-07-01 Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Departamento de Ciencia e Tecnologia do Ministerio da Saude (Decit/SCTIE/MS) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) BACKGROUND: Heart transplantation is a valuable therapeutic option for Chagas disease patients with severe cardiomyopathy. During patient follow-up, the differential diagnosis between cardiac transplant rejection and Chagas disease infection reactivation remains a challenging task, which hinders rapid implementation of the appropriate treatment. Herein we investigate whether polymerase chain reaction (PCR) strategies could facilitate early detection of Trypanosoma cruzi (T cruzi) in transplanted endomyocardial biopsies (EMBs). METHODS: In this study we analyzed 500 EMB specimens obtained from 58 chagasic cardiac transplant patients, using PCR approaches targeted to nuclear (rDNA 24S alpha) and kinetoplastid (kDNA) markers, and compared the efficiency of these approaches with that of other tests routinely used. RESULTS: T cruzi DNA was detected in 112 EMB specimens derived from 39 patients (67.2%). The first positive result occurred at a median 1.0 month post-transplant. Conventional histopathologic, blood smear and hemoculture analyses showed lower sensitivity and higher median time to the first positive result. Patient follow-up revealed that 31 of 39 PCR-positive cases presented clinical reactivation of Chagas disease at different time-points after transplantation. PCR techniques showed considerable sensitivity (0.82) and specificity (0.60), with area under the receiver operating characteristic (ROC) curves of 0.708 (p = 0.001). Moreover, PCR techniques anticipated the clinical signs of Chagas disease reactivation by up to 36 months, with a median time of 6 months and an average of 9.1 months. CONCLUSIONS: We found a good association between the PCR diagnosis and the clinical signs of the disease, indicating that the PCR approaches used herein are suitable for early diagnosis of Chagas disease reactivation, with high potential to assist physicians in treatment decisions. For this purpose, an algorithm is proposed for surveillance based on the molecular tests. (C) 2017 International Society for Heart and Lung Transplantation. All rights reserved. Univ Fed Minas Gerais, Inst Ciencias Biol, Dept Bioquim & Imunol, Ave Antonio Carlos 6627, BR-31270901 Belo Horizonte, MG, Brazil Univ Estadual Paulista, Inst Biociencias, Dept Genet, Sao Paulo, Brazil Univ Fed Minas Gerais, Hosp Clin, Belo Horizonte, MG, Brazil Univ Fed Minas Gerais, Fac Med, Dept Cirurgia, Belo Horizonte, MG, Brazil Univ Fed Minas Gerais, Inst Ciencias Biol, Dept Parasitol, Belo Horizonte, MG, Brazil Fiocruz Minas, Ctr Pesquisas Rene Rachou, Grp Integrado Pesquisas Biomarcadores, Belo Horizonte, MG, Brazil Univ Fed Minas Gerais, Fac Med, Dept Patol, Belo Horizonte, MG, Brazil Univ Fed Minas Gerais, Fac Med, Dept Clin Med, Belo Horizonte, MG, Brazil Univ Estadual Paulista, Inst Biociencias, Dept Genet, Sao Paulo, Brazil

Here we report the draft genome sequence of Metschnikowia australis strain UFMG-CM-Y6158, a yeast endemic to Antarctica. We isolated the strain from the marine seaweed Acrosiphonia arcta ( Chlorophyta ). The genome is 14.3 Mb long and contains 4,442 predicted protein-coding genes.

Trypanosoma cruzi is a protozoan parasite and the causative agent of Chagas disease. Like most living organisms, it is susceptible to oxidative stress, and must adapt to distinct environments. Hence, DNA repair is essential for its survival and the persistence of infection. Therefore, we studied whether T. cruzi has a homolog counterpart of the MutY enzyme (TcMYH), important in the DNA Base Excision Repair (BER) mechanism. Analysis of T. cruzi genome database showed that this parasite has a putative MutY DNA glycosylase sequence. We performed heterologous complementation assays using this genomic sequence. TcMYH complemented the Escherichia coli MutY- strain, reducing the mutation rate to a level similar to wild type. In in vitro assays, TcMYH was able to remove an adenine that was opposite to 8-oxoguanine. We have also constructed a T. cruzi lineage that overexpresses MYH. Although in standard conditions this lineage has similar growth to control cells, the overexpressor is more sensitive to hydrogen peroxide and glucose oxidase than the control, probably due to accumulation of AP sites in its DNA. Localization experiments with GFP-fused TcMYH showed this enzyme is present in both nucleus and mitochondrion. QPCR and MtOX results reinforce the presence and function of TcMYH in these two organelles. Our data suggest T. cruzi has a functional MYH DNA glycosylase, which participates in nuclear and mitochondrial DNA Base Excision Repair.

Despite their abundance, the molecular functions of long non-coding RNAs in mammalian nervous systems remain poorly understood. Here we show that the long non-coding RNA, NEAT1, directly modulates neuronal excitability and is associated with pathological seizure states. Specifically, NEAT1 is dynamically regulated by neuronal activity in vitro and in vivo, binds epilepsy-associated potassium channel-interacting proteins including KCNAB2 and KCNIP1, and induces a neuronal hyper-potentiation phenotype in iPSC-derived human cortical neurons following antisense oligonucleotide knockdown. Next generation sequencing reveals a strong association of NEAT1 with increased ion channel gene expression upon activation of iPSC-derived neurons following NEAT1 knockdown. Furthermore, we show that while NEAT1 is acutely down-regulated in response to neuronal activity, repeated stimulation results in NEAT1 becoming chronically unresponsive in independent in vivo rat model systems relevant to temporal lobe epilepsy. We extended previous studies showing increased NEAT1 expression in resected cortical tissue from high spiking regions of patients suffering from intractable seizures. Our results indicate a role for NEAT1 in modulating human neuronal activity and suggest a novel mechanistic link between an activity-dependent long non-coding RNA and epilepsy.

In recent years, proteasome involvement in the damage response induced by ionizing radiation (IR) became evident. However, whether proteasome plays a direct or indirect role in IR-induced damage response still unclear. Trypanosoma cruzi is a human parasite capable of remarkable high tolerance to IR, suggesting a highly efficient damage response system. Here, we investigate the role of T. cruzi proteasome in the damage response induced by IR. We exposed epimastigotes to high doses of gamma ray and we analyzed the expression and subcellular localization of several components of the ubiquitin-proteasome system. We show that proteasome inhibition increases IR-induced cell growth arrest and proteasome-mediated proteolysis is altered after parasite exposure. We observed nuclear accumulation of 19S and 20S proteasome subunits in response to IR treatments. Intriguingly, the dynamic of 19S particle nuclear accumulation was more similar to the dynamic observed for Rad51 nuclear translocation than the observed for 20S. In the other hand, 20S increase and nuclear translocation could be related with an increase of its regulator PA26 and high levels of proteasome-mediated proteolysis in vitro. The intersection between the opposed peaks of 19S and 20S protein levels was marked by nuclear accumulation of both 20S and 19S together with Ubiquitin, suggesting a role of ubiquitin-proteasome system in the nuclear protein turnover at the time. Our results revealed the importance of proteasome-mediated proteolysis in T. cruzi IR-induced damage response suggesting that proteasome is also involved in T. cruzi IR tolerance. Moreover, our data support the possible direct/signaling role of 19S in DNA damage repair. Based on these results, we speculate that spatial and temporal differences between the 19S particle and 20S proteasome controls proteasome multiple roles in IR damage response.