Pantelis Topalis

Co-transcriptional RNA-DNA hybrids can not only cause DNA damage threatening genome integrity but also regulate gene activity in a mechanism that remains unclear. Here, we show that the nucleotide excision repair factor XPF interacts with the insulator binding protein CTCF and the cohesin subunits SMC1A and SMC3, leading to R-loop–dependent DNA looping upon transcription activation. To facilitate R-loop processing, XPF interacts and recruits with TOP2B on active gene promoters, leading to double-strand break accumulation and the activation of a DNA damage response. Abrogation of TOP2B leads to the diminished recruitment of XPF, CTCF, and the cohesin subunits to promoters of actively transcribed genes and R-loops and the concurrent impairment of CTCF-mediated DNA looping. Together, our findings disclose an essential role for XPF with TOP2B and the CTCF/cohesin complex in R-loop processing for transcription activation with important ramifications for DNA repair–deficient syndromes associated with transcription-associated DNA damage.

Background: This study aimed to investigate the molecular profiles of 237 stage III CRC patients from the international IDEA study. It also sought to correlate these profiles with Toll-like and vitamin D receptor polymorphisms, clinicopathological and epidemiological characteristics, and patient outcomes. Methods: Whole Exome Sequencing and PCR-RFLP on surgical specimens and blood samples, respectively, were performed to identify molecular profiling and the presence of Toll-like and vitamin D polymorphisms. Bioinformatic analysis revealed mutational status. Results: Among the enrolled patients, 63.7% were male, 66.7% had left-sided tumors, and 55.7% received CAPOX as adjuvant chemotherapy. Whole exome sequencing identified 59 mutated genes in 11 different signaling pathways from the Kyoto Encyclopedia of Genes and Genomes (KEGG) CRC panel. On average, patients had 8 mutated genes (range, 2–21 genes). Mutations in ARAF and MAPK10 emerged as independent prognostic factors for reduced DFS (p = 0.027 and p < 0.001, respectively), while RAC3 and RHOA genes emerged as independent prognostic factors for reduced OS (p = 0.029 and p = 0.006, respectively). Right-sided tumors were also identified as independent prognostic factors for reduced DFS (p = 0.019) and OS (p = 0.043). Additionally, patients with tumors in the transverse colon had mutations in genes related to apoptosis, PIK3-Akt, Wnt, and MAPK signaling pathways. Conclusions: Molecular characterization of tumor cells can enhance our understanding of the disease course. Mutations may serve as promising prognostic biomarkers, offering improved treatment options. Confirming these findings will require larger patient cohorts and international collaborations to establish correlations between molecular profiling, clinicopathological and epidemiological characteristics and clinical outcomes.

Background: This study aimed to investigate the molecular profiles of stage III CRC patients from the international IDEA study. It also sought to correlate these profiles with Toll-like and vitamin D receptor polymorphisms, clinicopathological and epidemiological characteristics, and patient outcomes. Methods: Whole Exome Sequencing and PCR-RFLP on surgical specimens and blood samples, respectively, were performed to identify molecular profiling and the presence of Toll-like and vitamin D polymorphisms. Bioinformatic analysis revealed mutational status. Results: Among the enrolled patients, 63.7% were male, 66.7% had left-sided tumors, and 55.7% received CAPOX as adjuvant chemotherapy. Whole exome sequencing identified 59 mutated genes in 11 different signaling pathways from the Kyoto Encyclopedia of Genes and Genomes (KEGG) CRC panel. On average, patients had 8 mutated genes (range, 2-21 genes). Mutations in ARAF and MAPK10 emerged as independent prognostic factors for reduced DFS (p=0.027 and p<0.001, respectively), while RAC3 and RHOA genes emerged as independent prognostic factors for reduced OS (p=0.029 and p=0.006, respectively). Right-sided tumors were also identified as independent prognostic factors for reduced DFS (p=0.019) and OS (p=0.043). Additionally, patients with tumors in the transverse colon had mutations in genes related to apoptosis, PIK3-Akt, Wnt, and MAPK signaling pathways. Conclusions: Molecular characterization of tumor cells can enhance our understanding of the disease course. Mutations may serve as promising prognostic biomarkers, offering improved treatment options. Confirming these findings with require larger patient cohorts and international collaborations to establish correlations between molecular profiling, clinicopathological and epidemiological characteristics and clinical outcomes.

R-loops lead to chronic inflammation upon DNA damage.

RNA splicing, transcription and the DNA damage response are intriguingly linked in mammals but the underlying mechanisms remain poorly understood. Using an in vivo biotinylation tagging approach in mice, we show that the splicing factor XAB2 interacts with the core spliceosome and that it binds to spliceosomal U4 and U6 snRNAs and pre-mRNAs in developing livers. XAB2 depletion leads to aberrant intron retention, R-loop formation and DNA damage in cells. Studies in illudin S-treated cells and Csb^m/m developing livers reveal that transcription-blocking DNA lesions trigger the release of XAB2 from all RNA targets tested. Immunoprecipitation studies reveal that XAB2 interacts with ERCC1-XPF and XPG endonucleases outside nucleotide excision repair and that the trimeric protein complex binds RNA:DNA hybrids under conditions that favor the formation of R-loops. Thus, XAB2 functionally links the spliceosomal response to DNA damage with R-loop processing with important ramifications for transcription-coupled DNA repair disorders.

Only a small fraction of the mosquito species of the genus Anopheles are able to transmit malaria, one of the biggest killer diseases of poverty, which is mostly prevalent in the tropics. This diversity has genetic, yet unknown, causes. In a further attempt to contribute to the elucidation of these variances, the international “Anopheles Genomes Cluster Consortium” project (a.k.a. “16 Anopheles genomes project”) was established, aiming at a comprehensive genomic analysis of several anopheline species, most of which are malaria vectors. In the frame of the international consortium carrying out this project our team studied the genes encoding families of non-coding RNAs (ncRNAs), concentrating on four classes: microRNA (miRNA), ribosomal RNA (rRNA), small nuclear RNA (snRNA), and in particular small nucleolar RNA (snoRNA) and, finally, transfer RNA (tRNA). Our analysis was carried out using, exclusively, computational approaches, and evaluating both the primary NGS reads as well as the respective genome assemblies produced by the consortium and stored in VectorBase; moreover, the results of RNAseq surveys in cases in which these were available and meaningful were also accessed in order to obtain supplementary data, as were “pre-genomic era” sequence data stored in nucleic acid databases. The investigation included the identification and analysis, in most species studied, of ncRNA genes belonging to several families, as well as the analysis of the evolutionary relations of some of those genes in cross-comparisons to other members of the genus Anopheles. Our study led to the identification of members of these gene families in the majority of twenty different anopheline taxa. A set of tools for the study of the evolution and molecular biology of important disease vectors has, thus, been obtained.

Background With about half a billion cases, of which nearly one million fatal ones, malaria constitutes one of the major infectious diseases worldwide. A recently revived effort to eliminate the disease also focuses on IT resources for its efficient control, which prominently includes the control of the mosquito vectors that transmit the Plasmodium pathogens. As part of this effort, IDOMAL has been developed and it is continually being updated. Findings In addition to the improvement of IDOMAL’s structure and the correction of some inaccuracies, there were some major subdomain additions such as a section on natural products and remedies, and the import, from other, higher order ontologies, of several terms, which were merged with IDOMAL terms. Effort was put on rendering IDOMAL fully compatible as an extension of IDO, the Infectious Disease Ontology. The reason for the difficulties in fully reaching that target were the inherent differences between vector-borne diseases and “classical” infectious diseases, which make it necessary to specifically adjust the ontology’s architecture in order to comprise vectors and their populations. Conclusions In addition to a higher coverage of domain-specific terms and optimizing its usage by databases and decision-support systems, the new version of IDOMAL described here allows for more cross-talk between it and other ontologies, and in particular IDO. The malaria ontology is available for downloading at the OBO Foundry (http://www.obofoundry.org/cgi-bin/detail.cgi?id=malaria_ontology) and the NCBO BioPortal (http://bioportal.bioontology.org/ontologies/1311).

Genome sequencing of many eukaryotic pathogens and the volume of data available on public resources have created a clear requirement for a consistent vocabulary to describe the range of developmental forms of parasites. Consistent labeling of experimental data and external data, in databases and the literature, is essential for integration, cross database comparison, and knowledge discovery. The primary objective of this work was to develop a dynamic and controlled vocabulary that can be used for various parasites. The paper describes the Ontology for Parasite Lifecycle (OPL) and discusses its application in parasite research. The OPL is based on the Basic Formal Ontology (BFO) and follows the rules set by the OBO Foundry consortium. The first version of the OPL models complex life cycle stage details of a range of parasites, such as Trypanosoma sp., Leishmania sp., Plasmodium sp., and Shicstosoma sp. In addition, the ontology also models necessary contextual details, such as host information, vector information, and anatomical locations. OPL is primarily designed to serve as a reference ontology for parasite life cycle stages that can be used for database annotation purposes and in the lab for data integration or information retrieval as exemplified in the application section below. OPL is freely available at http://purl.obolibrary.org/obo/opl.owl and has been submitted to the BioPortal site of NCBO and to the OBO Foundry. We believe that database and phenotype annotations using OPL will help run fundamental queries on databases to know more about gene functions and to find intervention targets for various parasites. The OPL is under continuous development and new parasites and/or terms are being added.

Background: Ontologies are rapidly becoming a necessity for the design of efficient information technology tools, especially databases, because they permit the organization of stored data using logical rules and defined terms that are understood by both humans and machines. This has as consequence both an enhanced usage and interoperability of databases and related resources. It is hoped that IDOMAL, the ontology of malaria will prove a valuable instrument when implemented in both malaria research and control measures. Methods: The OBOEdit2 software was used for the construction of the ontology. IDOMAL is based on the Basic Formal Ontology (BFO) and follows the rules set by the OBO Foundry consortium. Results: The first version of the malaria ontology covers both clinical and epidemiological aspects of the disease, as well as disease and vector biology. IDOMAL is meant to later become the nucleation site for a much larger ontology of vector borne diseases, which will itself be an extension of a large ontology of infectious diseases (IDO). The latter is currently being developed in the frame of a large international collaborative effort. Conclusions: IDOMAL, already freely available in its first version, will form part of a suite of ontologies that will be used to drive IT tools and databases specifically constructed to help control malaria and, later, other vector-borne diseases. This suite already consists of the ontology described here as well as the one on insecticide resistance that has been available for some time. Additional components are being developed and introduced into IDOMAL.