Biology subjects

Molina, R.

Publications and source records attributed to Molina, R..

7 recordsLinked to original sources

bioRxiv2026

Post-translational toxin-antitoxin control and RecBCD surveillance underlie phage defense in a constitutively armed Type VI retron

Type VI retrons are bacterial defense systems that couple reverse transcription to phage immunity through dedicated toxin-antitoxin modules, yet the mechanisms linking ssDNA synthesis to toxin activation remain unclear. Here we characterize Retron-Sen3, a Type VI retron from Salmonella enterica encoding a small protein (SP) toxin, an HTH antitoxin, and a reverse transcriptase (RT). Unlike previously examined Type VI retrons, Retron-Sen3 constitutively produces RT-dependent ssDNA species in the absence of phage infection, and these levels remain unchanged during infection, establishing that defense activation is uncoupled from ssDNA synthesis. Comparative analyses revealed a conserved ncRNA architecture across the cl2075 lineage, including a previously undescribed stem-loop essential for defense, and an extended transcript encompassing the SP coding region within a structurally conserved RNA framework. SP is constitutively produced and intrinsically toxic, whereas HTH neutralizes SP toxicity, is required for basal viability, and physically associates with RT in vivo independently of catalytic activity; notably, the HTH-SP toxin-antitoxin module operates independently of RT and ncRNA. Phage escape mutants and genetic analyses establish that inhibition of the host RecBCD complex triggers defense activation, identifying RecBCD surveillance as the signal that couples phage infection to toxin activation. Together, these findings reveal a post-translational regulatory mechanism distinct from the msDNA-dependent translational control described for Retron-Vpa2, establishing mechanistic diversity within this retron family.

microbiology↗

bioRxiv2026

Are mosquito species present in Spain competent for Oropouche virus?

Oropouche virus (OROV; Orthobunyavirus) is an emerging arbovirus endemic to South America and the Caribbean, with imported cases in European countries, including Spain. Although primarily transmitted by biting midges (Culicoides spp.), OROV has been detected in several mosquito species, raising concerns about potential establishment in non-endemic regions. European populations of Aedes albopictus, and Culex pipiens, as well as the invasive Aedes aegypti, represent relevant models for assessing vector competence. Here, we evaluated vector competence of Spanish Cx. pipiens biotype molestus, Spanish Ae. albopictus, and Ae. aegypti (Liverpool strain), for the 2024 OROV outbreak strain. Female mosquitoes were orally exposed to infectious blood meals and maintained under controlled insectary conditions. In addition, an additional group received a second non-infectious blood meal. The survival of the mosquitoes was monitored, and infection, dissemination, and transmission rates were assessed at 7-, 14-, and 21-days post-infection. Vertical transmission of the virus to the progenies was also analyzed. Overall, Ae. albopictus exhibited low infection rates, with occasional dissemination and transmission events. Aedes aegypti and Cx. pipiens showed infection and occasional dissemination, but no evidence of transmission. A second non-infectious blood meal did not significantly affect infection, dissemination, or transmission rates in any species. Viral loads in bodies and legs were low and did not differ significantly between species, time points, or feeding regimens. Survival was not affected by infection or blood-feeding regime. We did not find vertical transmission of OROV to the progenies. Regardless of virus dissemination in mosquitoes, our study indicates poor vector competence of Spanish Ae. albopictus and a lack of competence in Ae. aegypti and Cx. pipiens for the circulating OROV strain. These findings suggest a low risk for local OROV establishment in Spain, although continuous surveillance and research are warranted to monitor potential vector-virus adaptation.

microbiology↗

bioRxiv2021

Voltage imaging in Drosophila using a hybrid chemical-genetic rhodamine voltage reporter

We combine a chemically-synthesized, voltage-sensitive fluorophore with a genetically encoded, self-labeling enzyme to enable voltage imaging in Drosophila melanogaster. Previously, we showed that a rhodamine voltage reporter (RhoVR) combined with the HaloTag self-labeling enzyme could be used to monitor membrane potential changes from mammalian neurons in culture and brain slice. Here, we apply this hybrid RhoVR-Halo approach in vivo to achieve selective neuron labeling in intact fly brains. We generate a Drosophila UAS-HaloTag reporter line in which the HaloTag enzyme is expressed on the surface of cells. We validate the voltage sensitivity of this new construct in cell culture before driving expression of HaloTag in specific brain neurons in flies. We show that selective labeling of synapses, cells, and brain regions can be achieved with RhoVR-Halo in either larval neuromuscular junction (NMJ) or in whole adult brains. Finally, we validate the voltage sensitivity of RhoVR-Halo in fly tissue via dual-electrode/imaging at the NMJ, show the efficacy of this approach for measuring synaptic excitatory post-synaptic potentials (EPSPs) in muscle cells, and perform voltage imaging of carbachol-evoked depolarization and osmolarity-evoked hyperpolarization in projection neurons and in interoceptive subesophageal zone neurons in fly brain explants following in vivo labeling. We envision the turn-on response to depolarizations, fast response kinetics, and two-photon compatibility of chemical indicators, coupled with the cellular and synaptic specificity of genetically-encoded enzymes, will make RhoVR-Halo a powerful complement to neurobiological imaging in Drosophila. Significance StatementVoltage imaging is a powerful method for interrogating neurobiology. Chemical indicators possess fast response kinetics, turn-on responses to membrane depolarization, and can be compatible with two-photon excitation. However, selective cell labeling in intact tissues and in vivo remains a challenge for completely synthetic fluorophores. Here, we show that a chemical - genetic hybrid approach in Drosophila enables cell-specific staining in vivo and voltage imaging in whole-brain explants and at neuromuscular junction synapses.

neuroscience↗

bioRxiv2021

Combining Attention-based Multiple Instance Learning and Gaussian Processes for CT Hemorrhage Detection

Intracranial hemorrhage (ICH) is a life-threatening emergency with high rates of mortality and morbidity. Rapid and accurate detection of ICH is crucial for patients to get a timely treatment. In order to achieve the automatic diagnosis of ICH, most deep learning models rely on huge amounts of slice labels for training. Unfortunately, the manual annotation of CT slices by radiologists is time-consuming and costly. To diagnose ICH, in this work, we propose to use an attention-based multiple instance learning (Att-MIL) approach implemented through the combination of an attention-based convolutional neural network (Att-CNN) and a variational Gaussian process for multiple instance learning (VGP-MIL). Only labels at scan-level are necessary for training. Our method trains the model using scan labels and assigns each slice with an attention weight, which can be used to provide slice-level predictions, and uses the VGPMIL model based on low-dimensional features extracted by the Att-CNN to obtain improved predictions both at slice and scan levels. To analyze the performance of the proposed approach, our model has been trained on 1150 scans from an RSNA dataset and evaluated on 490 scans from an external CQ500 dataset. Our method outperforms other methods using the same scan-level training and is able to achieve comparable or even better results than other methods relying on slicelevel annotations.

bioinformatics↗

bioRxiv2021

Comparative analysis of mammal genomes unveils key genomic variability for human lifespan.

Mammals vary 100-fold in their maximum lifespan. This enormous variation is the result of the adaptations of each species to their own biological trade-offs and ecological conditions. Comparative genomics studies have demonstrated that the genomic factors underlying the lifespans of species and the longevity of individuals are shared across the tree of life. Here, we set out to compare protein-coding regions across the mammalian phylogeny, aiming to detect individual amino acid changes shared by the most long-lived mammal species and genes whose rates of protein evolution correlate with longevity. We discovered a total of 2,737 amino acid changes in 2,004 genes that distinguish long- and short-lived mammals, significantly more than expected by chance (p=0.003). The detected genes belong to pathways involved in regulating lifespan, such as inflammatory response and hemostasis. Among them, a total 1,157 amino acids, located in 996 different genes, showed a significant association with maximum lifespan in a phylogenetically controlled test. Interestingly, most of the detected amino acids positions do not vary in extant human populations (>81.2%) or have allele frequencies below 1% (99.78%), Consequently, almost none could have been detected by Genome-Wide Association Studies (GWAS). Additionally, we identified four more genes whose rate of protein evolution correlated with longevity in mammals. Crucially, SNPs located in the detected genes explain a larger fraction of human lifespan heritability than expected by chance, successfully demonstrating for the first time that comparative genomics can be used to enhance the interpretation of human GWAS. Finally, we show that the human longevity-associated proteins coded by the detected genes are significantly more stable than the orthologous proteins from short-lived mammals, strongly suggesting that general protein stability is linked to increased lifespan.

genomics↗

bioRxiv2020

A collection of designed peptides to target SARS-Cov-2 - ACE2 interaction: PepI-Covid19 database.

The angiotensin-converting enzyme 2 is the cellular receptor used by SARS coronavirus SARS-CoV and SARS-CoV-2 to enter the cell. Both coronavirus use the receptor-binding domain (RBD) of their viral spike protein to interact with ACE2. The structural basis of these interactions are already known, forming a dimer of ACE2 with a trimer of the spike protein, opening the door to target them to prevent the infection. Here we present PepI-Cov19 database, a repository of peptides designed to target the interaction between the RDB of SARS-CoV-2 and ACE2 as well as the dimerization of ACE2 monomers. The peptides were modelled using our method PiPreD that uses native elements of the interaction between the targeted protein and cognate partner that are subsequently included in the designed peptides. These peptides recapitulate stretches of residues present in the native interface plus novel and highly diverse conformations that preserve the key interactions on the interface. PepI-Covid19 database provides an easy and convenient access to this wealth of information to the scientific community with the view of maximizing its potential impact in the development of novel therapeutic agents.

bioinformatics↗

bioRxiv2020

A Zinc-dependent metalloproteinase in the intracellular adaptation of Brucella abortus in macrophages

Brucella abortus is a pathogen that survives in macrophages. Several virulence factors participate in this process, including the open reading frame (ORF) BAB1_0270 codifying of a Zinc-dependent metalloproteinase. Here, its contribution in the process of intracellular adaptation was analyzed by infecting RAW264.7 macrophages with the mutant B. abortus {Delta}270 strain. Results showed that this Zinc-dependent metalloproteinase is a cytoplasmic protein that conforms an operon with a transcriptional regulator, which may constitute a type II toxin-antitoxin system. Functionally, this Zinc-dependent metalloproteinase participated neither in the adherence nor the initial intracellular traffic of B. abortus in macrophages. Nevertheless, its deletion significantly increased the co-localization of B. abortus {Delta}270 with phagolysosomal cathepsin D, reducing both its co-localization with calnexin, present in endoplasmic reticulum derived vesicles, and its intracellular replication within macrophages. Besides, B. abortus {Delta}270-infected macrophages produced significantly higher levels of TNF-, IL-6, CD80 and CD86 than B. abortus 2308, even when several genes involved in virulence (vjbR, hutC, bvrR, virB1) were up-regulated in this mutant. Finally, its deletion significantly reduced the capacity of B. abortus {Delta}270 to adapt, grow and express several virulence factors under acidic conditions. Based on these results, we discuss the role of this Zinc-dependent metalloproteinase in the regulation of the virulence of this pathogen, concluding that it contributes significantly to the intracellular adaptation of B. abortus 2308 during the infection of macrophages. Author summaryBrucella abortus is the causative agent of the brucellosis, a highly contagious diseases. A Zinc-dependent metalloproteinase contributes significantly in the intracellular survival. Here, we demonstrate that this metalloproteinase has homology with ImmA/IrrE proteases, which are involved in the bacterial resistance to hostile environment. Furthermore, it conforms a gene pair with a transcriptional regulator, being required by B. abortus to escape from phagolysosomes, to achieve the endoplasmic reticulum and replicate within macrophages. Its deletion from B. abortus stimulated the macrophages, which produced higher levels of pro-inflammatory cytokines and co-stimulatory proteins. This pathogen showed a reduced ability to adapt and grow under acidic conditions, which would negatively affect its escape from phagolysosomes and consequently, stimulating macrophages. Therefore, this work describes how this Zinc-dependent metalloproteinase significantly contributes in the intracellular adaptation of B. abortus 2308 in macrophages.

microbiology↗