Biology subjects

Enguita, F. J.

Publications and source records attributed to Enguita, F. J..

10 recordsLinked to original sources

bioRxiv2026

Verification of human nucleotide sequence reagents and cell line identities in original circRNA articles published in high impact factor journals

Numbers of research articles studying circRNAs have increased rapidly since 2017. Previous analyses of human circRNA articles in two high impact factor cancer research journals identified papers with wrongly identified nucleotide sequence reagents and circRNAs whose identities could not be independently verified. In the present study, verification of human nucleotide sequence reagent and cell line identities in retracted circRNA articles published from 2017-2021 in high impact factor journals found wrongly identified nucleotide sequences and/or cell lines in all 13 retracted papers. Similar analyses of human circRNA papers published in high impact factor journals in 2022 found wrongly identified, non-verifiable and/or questionable reagents in 71% (84/118) papers, where 51% (60/118) papers described at least one wrongly identified reagent. When individual error types and features of concern were considered, 2022 circRNA papers described wrongly identified nucleotide sequence reagents (52/118, 44%), questionable circRNA probes that did not meet accepted targeting requirements (34/118, 29%), non-verifiable nucleotide sequences (25/118, 21%), wrongly identified cell lines (22/118, 19%), and/or non-verifiable cell line identifiers (6/118, 5%). In summary, wrongly identified, non-verifiable and/or questionable reagents were unexpectedly frequent in human circRNA papers in high impact journals, highlighting the need for critical engagement with the circRNA literature.

genomics↗

bioRxiv2025

Dengue virus capsid protein interaction with nucleic acids

Dengue virus (DENV) transmission has greatly increased in the last decade, in part due to the geographical expansion of its vector, the Aedes spp. mosquitoes. These arthropods are now found even in temperate climates, including Europe, where outbreaks have already occurred. A better understanding of the virus life cycle is essential, as it may enable the development of specific treatments or therapies which are currently lacking. However, recent breakthroughs concerning the biologically relevant viral capsid (C) protein structure and function are encouraging. It is now clear the C protein binds both to host lipid droplets and to the viral genome, a set of interactions that is crucial for viral encapsidation and replication. Previously, we studied DENV C protein interaction with lipid droplets, at the molecular level. Here, we examine how the association of DENV C protein with the viral genome occurs. Using DENV C protein and single-stranded DNA (ssDNA) sequences analogous to previously identified important regions of the viral genome, we biophysically characterize their interactions. A decrease in fluorescence intensity and lifetime, an increase in both anisotropy and hydrodynamic diameter, as well as changes in the proteins secondary structure, were observed upon binding to ssDNA. These changes are consistent with molecular condensation and raise the possibility of liquid-liquid phase separation involvement in DENV C-nucleic acid complex formation. These findings may be useful for the development of future therapeutic strategies based on nucleic acids, not only for DENV but also for other related viruses.

biophysics↗

bioRxiv2024

PINK1-G411S mutant increases kinase stability and enhances mitochondrial-linked functions

PINK1, a mitochondria targeted Serine/Threonine kinase, regulates ATP production by phosphorylating the Complex I subunit NdufA10. However, when in the presence of depolarized mitochondria, PINK1 phosphorylates ubiquitin and Parkin triggering mitochondria clearance. Mutations in PINK1 have been linked to early-onset recessive familial forms of Parkinsons disease (PD). Deficits in Complex I enzymatic activity and an increase in oxidative damage have been identified in multiple brain regions of PD patients. Unravelling how PINK1 activity regulates mitochondria fate is pivotal. In the present study we characterized how human PD-related PINK1 mutants affect major PINK1 functions. Using molecular dynamics, we gain mechanistic insight into how specific mutations alter the tertiary structure and stability of PINK1s ATP-binding pocket, leading to an increased rigidity and stability. More importantly, we report a structural explanation for the enhanced kinase function of the PINK1-G411S mutant.

cell biology↗

bioRxiv2023

Verification of nucleotide sequence reagent identities in original publications in high impact factor cancer research journals

Human gene research studies that describe wrongly identified nucleotide sequence reagents have been mostly identified in journals of low to moderate impact factor, where unreliable findings could be considered to have limited influence on future research. This study examined whether papers describing wrongly identified nucleotide sequences are also published in high impact factor cancer research journals. We manually verified nucleotide sequence identities in original Molecular Cancer articles published in 2014, 2016, 2018 and 2020, including nucleotide sequence reagents that were claimed to target circRNAs. Using keywords identified in problematic 2018 and 2020 Molecular Cancer papers, we also verified nucleotide sequence identities in 2020 Oncogene papers that studied miRNA(s) and/or circRNA(s). Overall, 3.8% (253/6,647) and 4.3% (50/1,165) nucleotide sequences that were verified in Molecular Cancer and Oncogene papers, respectively, were found to be wrongly identified. These wrongly identified nucleotide sequences were distributed across 18% (92/500) original Molecular Cancer papers, including 38% Molecular Cancer papers from 2020, and 40% (21/52) selected Oncogene papers from 2020. Original papers with wrongly identified nucleotide sequences were therefore unexpectedly frequent in two high impact factor cancer research journals, highlighting the risks of employing journal impact factors or citations as proxies for research quality.

cancer biology↗

bioRxiv2022

Loss of the protein disrupted in schizophrenia 1 (DISC1) impairs cardiac resistance against ischemia.

Molecular targets that contribute to post-myocardial infarction heart failure remains elusive. Here we studied the global transcriptional landscape of human inducible pluripotent stem cell-derived cardiomyocytes under simulated ischemia at the whole genome level in an unbiased manner. Unexpectedly, we identified Disrupted in Schizophrenia 1 (DISC1), which has been almost exlusively studied in neurodevelopment/neurosignaling, as a key molecule. Reduced DISC1 levels increase infarct size in mouse hearts, increases cTnT release following surgery in coronary artery bypass grafting patients and decrease the survival of human AC16 cardiomyocytes exposed to ischemic conditions by disrupting cardioprotective pathways. Mechanistically, the loss of DISC1 interaction with Glycogen Synthase Kinase 3 Beta (GSK3{beta}) heightens GSK3{beta} activity, promoting cell death. Conversely, increasing DISC1 levels enhances cardioprotective signaling by maintaining the DISC1-GSK3{beta} interaction. Although DISC1 is known to interact with many proteins as an intracellular hub, we have identified that its specific interaction with GSK3{beta} is crucial for cardioprotection. These data also indicate that a key molecule involved in brain health and disease also contributes to cardiac health and disease, supporting the idea that neuropsychiatric conditions are systemic. SummaryThrough an unbiased screening at the whole genome level, we underscore that DISC1 is a key molecule for cardiomyocyte resilience post-myocardial infarction. DISC1 has been studied almost exclusively in neuroscience, involving in multiple cellular mechanisms. In contrast, we highlight DISC1s interaction with GSK3{beta} as specific mechanism that drives cardioprotection. We also report DISC1s implication in cardiac health following coronary artery bypass grafting.

molecular biology↗

bioRxiv2022

The interplay between lncRNAs, RNA-binding proteins and viral genome during SARS-CoV-2 infection reveals strong connections with regulatory events involved in RNA metabolism and immune response.

Viral infections are complex processes based on an intricate network of molecular interactions. The infectious agent hijacks components of the cellular machinery for its profit, circumventing the natural defense mechanisms triggered by the infected cell. The successful completion of the replicative viral cycle within a cell depends on the function of viral components versus the cellular defenses. Non-coding RNAs (ncRNAs) are important cellular modulators, either promoting or preventing the progression of viral infections. Among these ncRNAs, the long non-coding RNA (lncRNA) family is especially relevant due to their intrinsic functional properties and ubiquitous biological roles. Specific lncRNAs have been recently characterized as modulators of the cellular response during infection of human host cells by single stranded RNA viruses. However, the role of host lncRNAs in the infection by human RNA coronaviruses such as SARS-CoV-2 remains uncharacterized. In the present work, we have performed a transcriptomic study of a cohort of patients with different SARS-CoV-2 viral load. Our results revealed the existence of a SARS-CoV-2 infection-dependent pattern of transcriptional up-regulation in which specific lncRNAs are an integral component. To determine the role of these lncRNAs, we performed a functional correlation analysis complemented with the study of the validated interactions between lncRNAs and RNA-binding proteins (RBPs). This combination of in silico functional association studies and experimental evidence allowed us to identify a lncRNA signature composed of six elements - NRIR, BISPR, MIR155HG, FMR1-IT1, USP30-AS1, and U62317.2 - associated with the regulation of SARS-CoV-2 infection. We propose a competition mechanism between the viral RNA genome and the regulatory lncRNAs in the sequestering of specific RBPs that modulates the interferon response and the regulation of RNA surveillance by nonsense-mediated decay (NMD). O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=182 SRC="FIGDIR/small/485903v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@4368c9org.highwire.dtl.DTLVardef@1948201org.highwire.dtl.DTLVardef@e31fd9org.highwire.dtl.DTLVardef@1400805_HPS_FORMAT_FIGEXP M_FIG C_FIG Graphical abstractModel of interactions among lncRNA and cognate RNA-binding proteins in SARS-CoV-2 infection. According to our model, the viral genome can establish direct interactions with three core proteins (DDX3X, UPF1 and IGF2BP2) involved in mRNA metabolism and regulation of the interferon response, which are also components of a SARS-CoV-2 lncRNA-centered regulatory network. The competition between viral RNA and lncRNAs could act as a counteracting factor for the normal function of homeostatic lncRNA-centered regulatory networks, contributing to viral progression and replication. Black arrows depict physical interactions between network components; red arrows represent functional relationships.

bioinformatics↗

bioRxiv2021

The Great Deceiver: miR-2392's Hidden Role in Driving SARS-CoV-2 Infection

MicroRNAs (miRNAs) are small non-coding RNAs involved in post-transcriptional gene regulation that have a major impact on many diseases and provides an exciting avenue towards antiviral therapeutics. From patient transcriptomic data, we have discovered a circulating miRNA, miR-2392, that is directly involved with SARS-CoV-2 machinery during host infection. Specifically, we show that miR-2392 is key in driving downstream suppression of mitochondrial gene expression, increasing inflammation, glycolysis, and hypoxia as well as promoting many symptoms associated with COVID-19 infection. We demonstrate miR-2392 is present in the blood and urine of COVID-19 positive patients, but not detected in COVID-19 negative patients. These findings indicate the potential for developing a novel, minimally invasive, COVID-19 detection method. Lastly, using in vitro human and in vivo hamster models, we have developed a novel miRNA-based antiviral therapeutic that targets miR-2392, significantly reduces SARS-CoV-2 viability in hamsters and may potentially inhibit a COVID-19 disease state in humans.

systems biology↗

bioRxiv2020

Hsp27 reduces glycation-induced toxicity and aggregation of α-synuclein

-synuclein (aSyn) is a major player in Parkinsons disease (PD) and a group of other disorders collectively known as synucleinopathies, but the precise molecular mechanisms involved are still unclear. aSyn, as virtually all proteins, undergoes a series of posttranslational modifications (PTMs) during its lifetime which can affect its biology and pathobiology. We recently showed that glycation of aSyn by methylglyoxal (MGO) potentiates its oligomerization and toxicity, induces dopaminergic neuronal cell loss in mice, and affects motor performance in flies. Small heat-shock proteins (sHsps) are molecular chaperones that facilitate the folding of proteins or target misfolded proteins for clearance. Importantly, sHsps were shown to prevent aSyn aggregation and cytotoxicity. Upon treating cells with increasing amounts of methylglyoxal, we found that the levels of Hsp27 decreased in a dose-dependent manner. Therefore, we hypothesized that restoring the levels of Hsp27 in glycating environments could alleviate the pathogenicity of aSyn. Consistently, we found that Hsp27 reduced MGO-induced aSyn aggregation in cells, leading to the formation of non-toxic aSyn species. Remarkably, increasing the levels of Hsp27 suppressed the deleterious effects induced by MGO. Our findings suggest that in glycating environments, the levels of Hsp27 are important for modulating the glycation-associated cellular pathologies in synucleinopathies.

neuroscience↗

bioRxiv2019

Genome-wide Associations of Flavivirus Capsid Proteins

Dengue virus (DENV) and Zika virus (ZIKV) are both positive sense single-stranded RNA viruses. They are packaged within the virion with a capsid (C) protein to form the nucleocapsid. Based on cryo-electron microscopy imaging, the nucleocapsid has been described as lacking symmetry, whilst there is distinguishable separation of the C proteins from the viral RNA (vRNA) genome. Here, to elucidate the architecture of the nucleocapsid of DENV serotype 2 and ZIKV, we used a nuclease digestion assay and next-generation sequencing to map the respective vRNA genome wide association with the C protein in vitro. The C protein exhibited non-uniform binding along the vRNA, and as C protein concentration increased, the normalized read counts also increased. A saturation point of 1:100 (vRNA:C protein monomers) was found, and binding regions showed variable saturation patterns. We also observed that C protein had a preference for G-rich sequences for both viruses. Taken together, we demonstrate that the DENV 2 and ZIKV C proteins bind vRNA in a non-uniform manner with distinct patterns of association.\n\nSingificance StatementOur study demonstrates that flavivirus capsid proteins associate with the viral genome at specific sites rather than in a uniform manner as commonly expected. We estimate the number of capsid proteins binding to a single genomic RNA. We proceed to locate the capsid binding sites along the viral genomes of Dengue and Zika viruses. We characterize the binding sites in terms of affinity and analyze the nucleotide composition and sequence motifs at binding sites. We cross-reference binding sites against SHAPE reactivity data corresponding to local RNA secondary structure, which allows us to identify structural motifs of capsid binding sites. As capsid proteins are essential for viral packaging, these interactions may form attractive targets for therapeutic intervention.

genomics↗

bioRxiv2017

miRNAtools: advanced training using the miRNA web of knowledge

miRNAs are small non-coding RNAs, that act as negative regulators of the genomic output. Their intrinsic importance within cell biology and human disease is well known. Their mechanism of action based on the base pairing binding to their cognate targets, have helped the development of many computer applications for the prediction of miRNA target recognition. More recently, many other computer applications have appeared with the objective of studying miRNA roles in many contexts, trying to dissect and predict their functions in a specific biological process. Learning about miRNA function needs a practical training in the use of specific computer and web-based applications that are complementary to the wet-lab studies. In the last seven years we have been involved in the organization of advanced training courses about the in silico functional analysis of miRNAs and non-coding RNAs, for students ranging from the postgraduate to the post-doctoral level. In order to guide the learning process about miRNAs, we have created miRNAtools (http://mirnatools.eu), a web repository of miRNA tools and tutorials. This page is a compilation of tools to analyze miRNAs and their regulatory action that intends to collect and organize the information that is dispersed in the web. The miRNAtools webpage is completed by a collection of tutorials that can be used by students and tutors engaged in advanced training courses. The tutorials follow the rationale of the analysis of the function of selected miRNAs, starting from their nomenclature and genomic localization and finishing by assessing their involvement in specific cellular functions.\n\nDESCRIPTION OF THE AUTHORFrancisco J. Enguita is assistant professor at the Faculty of Medicine, University of Lisbon and senior investigator at the Instituto de Medicina Molecular within the same University.\n\nKEY POINTSO_LI- We have designed a web-page, mIRNAtools3 (http://mirnatools.eu) specifically devoted for advanced teaching purposes within the miRNA field.\nC_LIO_LI- The webpage constitutes also a repository of web-tools for the analysis of miRNA function in several contexts.\nC_LIO_LI- The webpage contains tutorials that cover many aspects related with miRNAs including nomenclature, target prediction and validation, and functional analysis.\nC_LI

bioinformatics↗