Papers by Julia Pinheiro Chagas Cunha
Additional file 5 of Replication origin location might contribute to genetic variability in Trypanosoma cruzi
Additional file 5. Distribution of ORIs among DGF-1 genes. A. Scatter plot of DGF-1 genes distrib... more Additional file 5. Distribution of ORIs among DGF-1 genes. A. Scatter plot of DGF-1 genes distribution according their gene size (in bp). Total DGF-1 and DGF-1 containing replication origins were compared regarding their gene size. *P-value
Additional file 4 of Replication origin location might contribute to genetic variability in Trypanosoma cruzi
Additional file 4. Schematic representation of a T. cruzi genomic region. CDSs are represented in... more Additional file 4. Schematic representation of a T. cruzi genomic region. CDSs are represented in green or purple vertical lines. Clusters of CDSs that are transcribed in the same direction represent a DGC. Arrows indicate the orientation of transcription. Green arrow is the transcription orientation of genes transcribed in positive strand (green) or in negative one (purple).
Additional file 2 of Replication origin location might contribute to genetic variability in Trypanosoma cruzi
Additional file 2. Distribution of consensus replication origin regions located at DGF-1 genes ac... more Additional file 2. Distribution of consensus replication origin regions located at DGF-1 genes according to their location at conserved (C- top) or disrupted (C-intermediate) T. cruzi genome domains (according to (43)). Both (B-below) represents DGF-1 containing origins that could be in either one domain. B. Examples of replication origins domains at conserved, disrupted and in between both domains. According to (43), DGF-1, RHS and GP63 (orange) may be located either at disrupted (red) or conserved (blue) compartments. The T. cruzi genome was loaded at UCSC genome browser and these 3 gene groups were colored in accordance. ORIs genome coordenates are shown in blue.
Approximately ten million people suffer from Chagas disease worldwide, caused by Trypanosoma cruz... more Approximately ten million people suffer from Chagas disease worldwide, caused by Trypanosoma cruzi, with the disease burden predominately focused in Latin America. Sleeping sickness is another serious health problem, caused by Trypanosoma brucei, especially in sub-Saharan countries. Unfortunately, the drugs currently available to treat these diseases have toxic effects and are not effective against all disease phases or parasite strains. Therefore, there is a clear need for the development of novel drugs and drug targets to treat these diseases. We propose the trypanosome prereplication machinery component, Orc1/Cdc6, as a potential target for drug development. In trypanosomes, Orc1/Cdc6 is involved in nuclear DNA replication, and, despite its involvement in such a conserved process, Orc1/Cdc6 is distinct from mammalian Orc1 and Cdc6 proteins. Moreover, RNAi-mediated silencing of trypanosome Orc1/Cdc6 expression in T. brucei decreased cell survival, indicating that Orc1/Cdc6 is crit...
HISTONAS EM Trypanosoma cruzi: SUAS VARIANTES E MODIFICAÇÕES AO LONGO DO CICLO DE VIDA
PLOS Pathogens, 2021
Trypanosoma cruzi alternates between replicative and nonreplicative life forms, accompanied by a ... more Trypanosoma cruzi alternates between replicative and nonreplicative life forms, accompanied by a shift in global transcription levels and by changes in the nuclear architecture, the chromatin proteome and histone posttranslational modifications. To gain further insights into the epigenetic regulation that accompanies life form changes, we performed genome-wide high-resolution nucleosome mapping using two T. cruzi life forms (epimastigotes and cellular trypomastigotes). By combining a powerful pipeline that allowed us to faithfully compare nucleosome positioning and occupancy, more than 125 thousand nucleosomes were mapped, and approximately 20% of them differed between replicative and nonreplicative forms. The nonreplicative forms have less dynamic nucleosomes, possibly reflecting their lower global transcription levels and DNA replication arrest. However, dynamic nucleosomes are enriched at nonreplicative regulatory transcription initiation regions and at multigenic family members,...
BackgroundGenomic organization and gene expression regulation in trypanosomes are remarkable beca... more BackgroundGenomic organization and gene expression regulation in trypanosomes are remarkable because protein-coding genes are organized into codirectional gene clusters with unrelated functions. Moreover, there is no dedicated promoter for each gene, resulting in polycistronic gene transcription, with posttranscriptional control playing a major role. Nonetheless, these parasites harbor epigenetic modifications at critical regulatory genome features that dynamically change among parasite stages, which are not fully understood.ResultsHere, we investigated the impact of chromatin changes in a scenario commanded by posttranscriptional control exploring the parasite Trypanosoma cruzi and its differentiation program using genome-wide approaches supported by transmission electron microscopy. The integration of FAIRE and MNase-seq data, two complementary epigenomic approaches, enabled us to identify differences in T. cruzi genome compartments, putative transcriptional start regions and viru...
FGF-2 induces a failure of cell cycle progression in cells harboring amplified K-Ras, revealing new insights into oncogene-induced senescence
Molecular Omics, 2021
Cells harboring oncogenic Ras were profiled with multi-omics to understand why they senesce inste... more Cells harboring oncogenic Ras were profiled with multi-omics to understand why they senesce instead of proliferate in response to growth factor signaling.
ABSTRACTParadoxically, oncogenes that drive cell cycle progression may also trigger pathways lead... more ABSTRACTParadoxically, oncogenes that drive cell cycle progression may also trigger pathways leading to senescence, thereby inhibiting the growth of tumorigenic cells. Along these lines, Y1 cells, which carry an amplification of Ras, become senescent after treatment with the mitogen FGF-2. To understand how FGF-2 promotes senescence, we profiled the epigenome, transcriptome, proteome, and phospho-proteome of Y1 cells stimulated with FGF-2. FGF-2 caused delayed acetylation of histone H4 and higher levels of H3K27me3. Sequencing analysis revealed decreased expression of cell cycle-related genes with concomitant loss of H3K27ac. In contrast, FGF-2 promoted the expression of p21, various cytokines, and MAPK-related genes. Nuclear envelope proteins, particularly lamin B1, displayed increased phosphorylation in response to FGF-2. Proteome analysis suggested alterations in cellular metabolism, as evident by modulated expression of enzymes involved in purine biosynthesis, tRNA aminoacylatio...
Genes, 2020
Trypanosoma cruzi is the etiological agent of Chagas disease, which affects millions of people in... more Trypanosoma cruzi is the etiological agent of Chagas disease, which affects millions of people in Latin America. No transcriptional control of gene expression has been demonstrated in this organism, and 50% of its genome consists of repetitive elements and members of multigenic families. In this study, we applied a novel bioinformatics approach to predict new repetitive elements in the genome sequence of T. cruzi. A new repetitive sequence measuring 241 nt was identified and found to be interspersed along the genome sequence from strains of different DTUs. This new repeat was mostly on intergenic regions, and upstream and downstream regions of the 241 nt repeat were enriched in surface protein genes. RNAseq analysis revealed that the repeat was part of processed mRNAs and was predominantly found in the 3′ untranslated regions (UTRs) of genes of multigenic families encoding surface proteins. Moreover, we detected a correlation between the presence of the repeat in the 3′UTR of multig...
BMC Genomics, 2020
Background: DNA replication in trypanosomatids operates in a uniquely challenging environment, si... more Background: DNA replication in trypanosomatids operates in a uniquely challenging environment, since most of their genomes are constitutively transcribed. Trypanosoma cruzi, the etiological agent of Chagas disease, presents high variability in both chromosomes size and copy number among strains, though the underlying mechanisms are unknown. Results: Here we have mapped sites of DNA replication initiation across the T. cruzi genome using Marker Frequency Analysis, which has previously only been deployed in two related trypanosomatids. The putative origins identified in T. cruzi show a notable enrichment of GC content, a preferential position at subtelomeric regions, coinciding with genes transcribed towards the telomeres, and a pronounced enrichment within coding DNA sequences, most notably in genes from the Dispersed Gene Family 1 (DGF-1). Conclusions: These findings suggest a scenario where collisions between DNA replication and transcription are frequent, leading to increased genetic variability, as seen by the increase SNP levels at chromosome subtelomeres and in DGF-1 genes containing putative origins.
Genome Research, 2019
DNA and histone proteins define the structure and composition of chromatin. Histone posttranslati... more DNA and histone proteins define the structure and composition of chromatin. Histone posttranslational modifications (PTMs) are covalent chemical groups capable of modeling chromatin accessibility, mostly due to their ability in recruiting enzymes responsible for DNA readout and remodeling. Mass spectrometry (MS)-based proteomics is the methodology of choice for large-scale identification and quantification of protein PTMs, including histones. High sensitivity proteomics requires online MS coupling with relatively low throughput and poorly robust nano-liquid chromatography (nanoLC) and, for histone proteins, a 2-d sample preparation that includes histone purification, derivatization, and digestion. We present a new protocol that achieves quantitative data on about 200 histone PTMs from tissue or cell lines in 7 h from start to finish. This protocol includes 4 h of histone extraction, 3 h of derivatization and digestion, and only 1 min of MS analysis via direct injection (DI-MS). We d...
Trichostatin A induces Trypanosoma cruzi histone and tubulin acetylation: effects on cell division and microtubule cytoskeleton remodelling
Parasitology, 2018
Trypanosoma cruzi, the causative agent of Chagas disease, is a public health concern in Latin Ame... more Trypanosoma cruzi, the causative agent of Chagas disease, is a public health concern in Latin America. Epigenetic events, such as histone acetylation, affect DNA topology, replication and gene expression. Histone deacetylases (HDACs) are involved in chromatin compaction and post-translational modifications of cytoplasmic proteins, such as tubulin. HDAC inhibitors, like trichostatin A (TSA), inhibit tumour cell proliferation and promotes ultrastructural modifications. In the present study, TSA effects on cell proliferation, viability, cell cycle and ultrastructure were evaluated, as well as on histone acetylation and tubulin expression of the T. cruzi epimastigote form. Protozoa proliferation and viability were reduced after treatment with TSA. Quantitative proteomic analyses revealed an increase in histone acetylation after 72 h of TSA treatment. Surprisingly, results obtained by different microscopy methodologies indicate that TSA does not affect chromatin compaction, but alters mi...
Frontiers in microbiology, 2018
Sporotrichosis is a mycosis that affects the skin, lymphatic system and other organs in humans an... more Sporotrichosis is a mycosis that affects the skin, lymphatic system and other organs in humans and animals. The disease has a worldwide distribution, with endemic areas in Brazil, and is caused by a complex of species, including . Some fungi release extracellular vesicles (EVs) that can interact with the host cell and modulate the host immune response. The aim of this study was to analyze the participation of EVs in the modulation of dendritic cells (DCs) and in the control of infection . Our results showed that , the EVs isolated from induced an increase in the phagocytic index and fungal burden in DCs. In addition, we observed a significant increase in IL-12p40 and TNF-α cytokine production. Then, the EVs were inoculated into BALB/c mice before subcutaneous infection with yeast, and the lesion was analyzed after 21, 35, and 42 days. An increase in fungal burden and lesion diameter were observed after 21 days in mice inoculated with a high concentration of EVs. However, after 35 da...
FGF-2 Antiproliferative Stimulation Induces Proteomic Dynamic Changes and High Expression of fosB and junB in K-Ras-Driven Mouse Tumor Cells
Proteomics, Jan 23, 2018
FGF-2 is a well-known cell proliferation promoter; however, it can also induce cell cycle arrest.... more FGF-2 is a well-known cell proliferation promoter; however, it can also induce cell cycle arrest. To gain insight into the molecular mechanisms of this antiproliferative effect, we investigated, for the first time, the early systemic proteomic differences induced by this growth factor in a K-Ras-driven mouse tumor cell line using a quantitative proteomics approach. More than 2900 proteins were quantified, indicating that terms associated with metabolism, RNA processing, replication, and transcription were enriched among proteins differentially expressed upon FGF2 stimulation. Proteomic trend dynamics indicated that, for proteins associated mainly with DNA replication and carbohydrate metabolism, an FGF2 stimulus delayed their abundance changes, whereas FGF2 stimulation accelerated other metabolic programs. Transcription regulatory network analysis indicated master regulators of FGF2 stimulation, including two critical transcription factors, fosB and junB. We investigated their expre...
Molecular & Cellular Proteomics, 2016
Chromatin associated proteins are key regulators of many important processes in the cell. Trypano... more Chromatin associated proteins are key regulators of many important processes in the cell. Trypanosoma cruzi, a protozoa flagellate that causes Chagas disease, alternates between replicative and nonreplicative forms accompanied by a shift on global transcription levels and by changes in its chromatin architecture. Here, we investigated the T. cruzi chromatin proteome using three different protocols and compared it between replicative (epimastigote) and nonreplicative (trypomastigote) forms by high-resolution mass spectrometry. More than 2000 proteins were identified and quantified both in chromatin and nonchromatin extracts. Besides histones and other known nuclear proteins, trypanosomes chromatin also contains metabolic (mainly from carbohydrate pathway), cytoskeleton and many other proteins with unknown functions. Strikingly, the two parasite forms differ greatly regarding their chromatin-associated factors composition and amount. Although the nucleosome content is the same for both life forms (as seen by MNase digestion), the remaining proteins were much less detected in nonreplicative forms, suggesting that they have a naked chromatin. Proteins associated to DNA proliferation, such as PCNA, RPA, and DNA topoisomerases were exclusively found in the chromatin of replicative stages. On the other hand, the nonreplicative stages have an enrichment of a histone H2B variant. Furthermore, almost 20% of replicative stages chromatin-associated proteins are expressed in nonreplicative forms, but located at nonchromatin space. We identified different classes of proteins including phosphatases and a Ran-binding protein, that may shuttle between chromatin and nonchromatin space during differentiation. Seven proteins, including those with unknown functions, were selected for further validation. We confirmed their location in chromatin and their differential expression, using Western blotting assays and chromatin immunoprecipitation (ChIP). Our results indicate that the replicative state in trypanosomes involves an increase of chromatin associated proteins content. We discuss in details, the qualitative and quantitative implication of this chromatin set in trypanosome chromatin biology. Because trypanosomes are early-branching organisms, this data can boost our understanding of chromatin-associated processes in other cell types.
Journal of proteome research, Jun 25, 2016
Histones are well-conserved proteins that form the basic structure of chromatin in eukaryotes and... more Histones are well-conserved proteins that form the basic structure of chromatin in eukaryotes and undergo several post-translational modifications, which are important for the control of transcription, replication, DNA damage repair and chromosome condensation. In early-branched organisms, histones are less conserved and appear to contain alternative sites for modifications, which could reveal evolutionary unique functions of histone modifications in gene expression and other chromatin-based processes. Here, by using high-resolution mass spectrometry, we identified and quantified histone post-translational modifications in two life cycle stages of Trypanosoma cruzi, the protozoan parasite that causes Chagas disease. We detected 44 new modifications, namely: 18 acetylations, 7 monomethylations, 8 dimethylations, 7 trimethylations and 4 phosphorylations. We found that replicative (epimastigote stage) contains more histone modifications than non-replicative and infective parasites (try...
Nuclear DNA replication in Trypanosomatid Protozoa
The parasites belonging to the family Trypanosomatidae (order Kinetoplastida) are among the most ... more The parasites belonging to the family Trypanosomatidae (order Kinetoplastida) are among the most primitive eukaryotes. Some trypanosomatids are the etiologic agents of neglected human pathologies such as South American and African trypanosomiasis and leishmaniasis. As a consequence of their ancient phylogenetic position, nuclear DNA replication in trypanosomatid protozoa shows conserved and non-conserved features. DNA replication in trypanosomatids initiates nearly simultaneously in the nucleus and in the genetic material of the single mitochondrion (or kinetoplast), suggesting that DNA synthesis is coordinately regulated in both organelles. In eukaryotes, nuclear DNA replication is preceded by assembly of the pre-replication complex, which is coordinated by the Origen Recognition Complex (ORC). However, in trypanosomatids, the prereplication complex differs from other eukaryotes and is similar to Archaea. All of these parasites contain only one protein that recognizes the replicati...
Cancer immunity, 2012
Cell surface proteins (CSPs) are excellent targets for the development of diagnostic and therapeu... more Cell surface proteins (CSPs) are excellent targets for the development of diagnostic and therapeutic reagents, and it is estimated that 10-20% of all genes in the human genome encode CSPs. In an effort to integrate all data publicly available for genes encoding cell surface proteins, a database (SurfaceomeDB) was developed. SurfaceomeDB is a gene-centered portal containing different types of information, including annotation for gene expression, protein domains, somatic mutations in cancer, and protein-protein interactions for all human genes encoding CSPs. SurfaceomeDB was implemented as an integrative and relational database in a user-friendly web interface, where users can search for gene name, gene annotation, or keywords. There is also a streamlined graphical representation of all data provided and links to the most important data repositories and databases, such as NCBI, UCSC Genome Browser, and EBI.
BioMed Research International, 2013
Introduction. Cell surface proteins are ideal targets for cancer therapy and diagnosis. We have i... more Introduction. Cell surface proteins are ideal targets for cancer therapy and diagnosis. We have identified a set of more than 3700 genes that code for transmembrane proteins believed to be at human cell surface.Methods. We used a high-throuput qPCR system for the analysis of 573 cell surface protein-coding genes in 12 primary breast tumors, 8 breast cell lines, and 21 normal human tissues including breast. To better understand the role of these genes in breast tumors, we used a series of bioinformatics strategies to integrates different type, of the datasets, such as KEGG, protein-protein interaction databases, ONCOMINE, and data from, literature.Results. We found that at least 77 genes are overexpressed in breast primary tumors while at least 2 of them have also a restricted expression pattern in normal tissues. We found common signaling pathways that may be regulated in breast tumors through the overexpression of these cell surface protein-coding genes. Furthermore, a comparison w...
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Papers by Julia Pinheiro Chagas Cunha