Biology subjects

Bellini, N. K.

Publications and source records attributed to Bellini, N. K..

3 recordsLinked to original sources

bioRxiv2025

First Genome-Wide Centromere Map of Trypanosoma cruzi Reveals Linear and 3D Compartment Boundaries and Spatial Clustering

Background: Trypanosoma cruzi, the etiological agent of Chagas disease, possesses a highly repetitive genome that has historically hindered high-quality assembly and structural characterization. Despite significant advances in assembling T. cruzi genomes, major gaps remain. Among these, the complete repertoire of centromeric sequences has remained elusive, representing a critical missing piece in our understanding of chromosome structure and inheritance. Results: Here, we generated high-coverage Hi-C (genome-wide chromosome conformation capture) data for the widely used T. cruzi Dm28c strain improving its genome assembly, reducing the number of scaffolds and producing a more contiguous and accurate genome. To investigate centromere organization, we performed ChIP-seq using the mNeonGreen-myc-tagged kinetochore proteins KKT2 and KKT3, resulting in the identification of 40 KKT-enriched peaks across 29 scaffolds. These peaks were located in regions enriched in retrotransposable elements, particularly L1Tc and VIPER, near strand switch regions, areas of high GC content, and at the boundaries between conserved genes and virulence-factor multigene families. Conclusion: Notably, Hi-C analysis revealed that centromeres may act as structural boundaries contributing to genome compartmentalization and frequently engage in 3D spatial clustering, suggesting a role in higher-order nuclear architecture. Overall, our study provides a high-quality reference genome for the Dm28c strain, presents the first genome-wide centromere map in T. cruzi, and offers novel insights into centromere-mediated 3D genome organization

genomics↗

bioRxiv2024

Hidden origami in Trypanosoma cruzi nuclei highlights its nonrandom 3D genomic organization

The protozoan Trypanosoma cruzi, the causative agent of Chagas disease, exhibits polycistronic transcription and unidimensional genome compartmentalization of core (conserved) and disruptive (virulence factors from multigenic families) genes. Approximately 50% of its genome is repetitive, mainly virulence factor genes. Genomic sequences, including repeats, motifs of architectural proteins, and noncoding RNA loci are crucial for genome folding. Here, we evaluated the genomic features associated with higher-order chromatin organization in T. cruzi through extensive computational processing of high-throughput chromosome conformation capture (Hi-C) data, accounting for repetitive regions and improvements in genome annotation. Our study revealed that repetitive DNA (multimapped reads) influences 3D chromatin folding, particularly in determining the boundaries of topologically associated domains (TAD)-like structures. Virulence factor genes, unlike core genes, form shorter and more compact TAD-like structures enriched in loops, suggesting a gene expression regulatory mechanism. We found nonprotein-coding RNA loci (e.g., tRNAs) and transcription termination sites preferentially located at the boundaries of the TAD-like structures, while pseudogenes and multigenic family genes located in unstructured genomic regions. Our data indicate 3D clustering of tRNA loci, likely optimizing transcription by RNA polymerase III, and a complex interaction between spliced-leader RNA and 18S rRNA loci. Our findings provide insights into 3D genome organization in T. cruzi, contributing to the understanding of supranucleosome-level chromatin organization and suggesting possible links between 3D architecture and gene expression. We draw an analogy to the art of origami (e.g., papers folded into various shapes) resembling the DNA packed in chromatin fibers assuming distinct folds within the nucleus. ImportanceDespite the knowledge about the linear genome sequence and the identification of numerous virulence factors in the protozoan parasite Trypanosoma cruzi, there has been a limited understanding of how these genomic features are spatially organized within the nucleus and how this organization impacts gene regulation and pathogenicity. By providing a detailed analysis of the three-dimensional chromatin architecture in T. cruzi, our study contributed to filling this gap. We deciphered part of the origami structure hidden in the T. cruzi nucleus, showing the unidimensional genomic features are nonrandomly organized in the nuclear 3D landscape. We revealed the possible role of non-protein-coding RNA loci (e.g., tRNAs, SL-RNA, and 18S RNA) in shaping the genomic architecture. These findings provide insights into an additional epigenetic layer that may influence gene expression. Graphical abstractThe spatial organization of chromatin within the nuclei of T. cruzi and its resemblance to origami art. A. Identification of the 3D nuclear architectures within T. cruzi nuclei: topologically associating domains (TADs) and their boundaries; chromatin loops; and 3D networks. Inter- and intrachromosomal interactions reflect DNA-DNA contacts on the same (cis) and between different (trans) chromosomes. B. Resemblance between origami art and chromatin folding. Steps "a" to "l" show the process of folding a flat piece of paper from its unidimensional view up to its 3D boat form. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/601582v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@1ca38b4org.highwire.dtl.DTLVardef@150bc99org.highwire.dtl.DTLVardef@18de8a9org.highwire.dtl.DTLVardef@1a5efa4_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2020

Trypanosomatid selenophosphate synthetase structure, function and interaction with selenocysteine lyase

Early branching eukaryotes have been used as models to study the evolution of cellular molecular processes. Strikingly, human parasite of the Trypanosomatidae family (T. brucei, T. cruzi and L. major) conserve the complex machinery responsible for selenocysteine biosynthesis and incorporation in selenoproteins (SELENOK/SelK, SELENOT/SelT and SELENOTryp/SelTryp), although these proteins do not seem to be essential for parasite viability under laboratory controlled conditions. Selenophosphate synthetase (SEPHS/SPS) plays an indispensable role in selenium metabolism, being responsible for catalyzing the formation of selenophosphate, the biological selenium donor for selenocysteine synthesis. We solved the crystal structure of the L. major selenophosphate synthetase and confirmed that its dimeric organization is functionally important throughout the domains of life. We also demonstrated its interaction with selenocysteine lyase (SCLY) and showed that it is not present in other stable complexes involved in the selenocysteine pathway, namely the phosphoseryl-tRNASec kinase (PSTK)-Sec-tRNASec synthase (SEPSECS) and the tRNASec-specific elongation factor (eEFSec)-ribosome. Endoplasmic reticulum stress with ditiothreitol (DTT) or tunicamycin upon selenophosphate synthetase ablation in procyclic T. brucei cells led to a growth defect. On the other hand, only DTT presented a negative effect in bloodstream T. brucei expressing selenophosphate synthetase-RNAi. Although selenoprotein T (SELENOT) was dispensable for both forms of the parasite, SELENOT-RNAi procyclic T. brucei cells were sensitive to DTT. Together, our data suggest a role for the T. brucei selenophosphate synthetase in regulation of the parasites ER stress response. SynopsisSelenium is both a toxic compound and a micronutrient. As a micronutrient, it participates in the synthesis of specific proteins, selenoproteins, as the amino acid selenocysteine. The synthesis of selenocysteine is present in organisms ranging from bacteria to humans. The protozoa parasites of the Trypanosomatidae family, that cause major tropical diseases, conserve the complex machinery responsible for selenocysteine biosynthesis and incorporation in selenoproteins. However, this pathway has been considered dispensable for the protozoa cells. This has intrigued us, and lead to question that if maintained in the cell it should be under selective pressure and therefore be necessary. Also, since the intermediate products of selenocysteine synthesis are toxic to the cell, it has been proposed that these compounds need to be sequestered from the cytoplasm. Therefore, extensive and dynamic protein-protein interactions must happen to deliver those intermediates along the pathway. In this study we have investigated the molecular and structural interactions of different proteins involved in selenocystein synthesis and describe its involvement in the endoplasmic reticulum protection to oxidative stress. Our results also show how the interaction of different proteins leads to the protection of the cell against the toxic effects of seleium compounds during selenocysteine synthesis.

molecular biology↗