Biology subjects

Perez, L.

Publications and source records attributed to Perez, L..

31 records · Page 2Linked to original sources

bioRxiv2022

Epigenetic biomarkers of autoimmune risk and protective antioxidant signaling in methylmercury-exposed adults

I.BackgroundEpigenome-wide association studies (EWAS) are a highly promising approach that can inform precision environmental health. However, current EWAS are underpowered for biomarker detection and increasing sample sizes will require substantial resources. Therefore, alternative approaches for identifying candidate biomarkers through EWAS are critical for moving the field forward. ObjectivesTo provide proof-of-principle that maximizing exposure variance in EWAS by selecting participants from disproportionately exposed global populations enables effective candidate biomarker detection, even in small sample sizes. MethodsWe profiled genome-wide DNA methylation using Illumina Infinium MethylationEPIC BeadChip in whole blood from N=32 individuals from Madre de Dios, Peru with high methylmercury (MeHg) exposure due to artisanal and small-scale gold mining. We compared DNA methylation in N=16 individuals with high (>10 g/g) vs. N=16 individuals with low (<1 g/g) total hair mercury (a proxy for methylmercury exposure), matched on age and sex. ResultsWe identified nine differentially methylated CpG sites (FDR<0.05), including several with known links to MeHg toxicity. The most significantly different CpG site was in an intronic enhancer of the SLC5A7 gene, which encodes the L-type amino acid transporter 1 (LAT1) that facilitates MeHg transport into protein-rich tissue, including muscle and brain. Our Gene Ontology and transcription factor motif enrichment analyses identified differential methylation of genes involved in several outcomes with established links to MeHg, including immune response, neurotoxicity, and type 2 diabetes (T2D) risk. Last, we identified candidate epigenetic biomarkers of PUFA-mediated protection against MeHg toxicity. DiscussionHere, we show that a small EWAS on samples with high MeHg exposure variance can detect candidate differentially methylated CpGs and pathways of interest relevant to MeHg biology. Similar EWAS in global populations with known high exposure variance can be leveraged to develop targeted, custom sequencing panels and microarrays limited to replicated, validated biomarkers of a given exposure.

genomics↗

bioRxiv2022

SARS-CoV-2 Omicron potently neutralized by a novel antibody with unique Spike binding properties

The SARS-CoV-2 Omicron variant exhibits very high levels of transmission, pronounced resistance to authorized therapeutic human monoclonal antibodies and reduced sensitivity to vaccine-induced immunity. Here we describe P2G3, a human monoclonal antibody (mAb) isolated from a previously infected and vaccinated donor, which displays picomolar-range neutralizing activity against Omicron BA.1, BA.1.1, BA.2 and all other current variants, and is thus markedly more potent than all authorized or clinically advanced anti-SARS-CoV-2 mAbs. Structural characterization of P2G3 Fab in complex with the Omicron Spike demonstrates unique binding properties to both down and up spike trimer conformations at an epitope that partially overlaps with the receptor-binding domain (RBD), yet is distinct from those bound by all other characterized mAbs. This distinct epitope and angle of attack allows P2G3 to overcome all the Omicron mutations abolishing or impairing neutralization by other anti-SARS-COV-2 mAbs, and P2G3 accordingly confers complete prophylactic protection in the SARS-CoV-2 Omicron monkey challenge model. Finally, although we could isolate in vitro SARS-CoV2 mutants escaping neutralization by P2G3 or by P5C3, a previously described broadly active Class 1 mAb, we found these viruses to be lowly infectious and their key mutations extremely rare in the wild, and we could demonstrate that P2G3/P5C3 efficiently cross-neutralized one anothers escapees. We conclude that this combination of mAbs has great prospects in both the prophylactic and therapeutic settings to protect from Omicron and other VOCs.

immunology↗

bioRxiv2021

Evolutionary analyses of visual opsin genes in anurans reveals diversity and positive selection suggestive of functional adaptation to distinct light environments

Among major vertebrate groups, anurans (frogs and toads) are understudied with regards to their visual systems and little is known about variation among species that differ in ecology. We sampled North American anurans representing diverse evolutionary and life histories that likely possess visual systems adapted to meet different ecological needs. Using standard molecular techniques, visual opsin genes, which encode the protein component of visual pigments, were obtained from anuran retinas. Additionally, we extracted the visual opsins from publicly available genome and transcriptome assemblies, further increasing the phylogenetic and ecological diversity of our dataset. We found that anurans consistently express four visual opsin genes (RH1, LWS, SWS1, and SWS2, but not RH2) even though reported photoreceptor complements vary widely among species. We found the first evidence of visual opsin duplication in an amphibian with the duplication of the LWS gene in the African bullfrog, which had distinct LWS copies on the sex chromosomes. The proteins encoded by these genes showed considerable sequence variation among species, including at sites known to shift the spectral sensitivity of visual pigments in other vertebrates and thus mediate dim-light and color vision. Using molecular evolutionary analyses of selection (dN/dS) we found significant evidence for positive selection at a subset of sites in the dim-light rod opsin gene RH1 and the long wavelength sensitive cone opsin gene LWS. The function of sites inferred to be under positive selection are largely unknown, but a few are likely to affect spectral sensitivity and other visual pigment functions based on proximity to previously identified sites in other vertebrates. The observed variation cannot fully be explained by evolutionary relationships among species alone. Taken together, our results suggest that other ecological factors, such as habitat and life history, as well as behaviour, may be driving changes to anuran visual systems.

evolutionary biology↗

bioRxiv2021

Cuban history of CRF19 recombinant subtype of HIV-1

CRF19 is a recombinant form of HIV-1 subtypes D, A1 and G, which was first sampled in Cuba in 1999, but was already present there in 1980s. CRF19 was reported almost uniquely in Cuba, where it accounts for ~25% of new HIV-positive patients and causes rapid progression to AIDS (~ 3 years). We analyzed a large data set comprising ~ 350 pol and env sequences sampled in Cuba over the last 15 years and ~ 350 from Los Alamos database. This data set contained both CRF19 (~ 315), and A1, D and G sequences. We performed and combined analyses for the three A1, G and D regions, using fast maximum likelihood approaches, including: (1) phylogeny reconstruction, (2) spatio-temporal analysis of the virus spread, and ancestral character reconstruction for (3) transmission mode and (4) drug resistance mutations (DRMs). This allowed us to acquire new insights on the CRF19 origin and transmission patterns. We showed that CRF19 recombined between 1966 and 1977, most likely in Cuban community stationed in Congo region. We further investigated CRF19 spread on the Cuban province level, and discovered that the epidemic started in 1970s, most probably in Villa Clara, that it was at first carried by heterosexual transmissions, and then quickly spread in the 1980s within the "men having sex with men" (MSM) community, with multiple transmissions back to heterosexuals. The analysis of the transmission patterns of common DRMs showed mostly acquired drug resistance rather than transmitted one. Our results show a very early introduction of CRF19 in Cuba, which could explain its local epidemiological success. Ignited by a major founder event, the epidemic then followed a similar pattern as other subtypes and CRFs in Cuba. The reason for the short time to AIDS remains to be understood and requires specific surveillance, in Cuba and elsewhere. Author summaryCRF19 is a recombinant form of HIV-1, which causes rapid progression to AIDS (~ 3 years versus 5 - 10 years for other subtypes and CRFs). CRF19 is reported almost uniquely in Cuba, where it is highly prevalent (~ 25%) among newly detected HIV-1 patients. In this study, we found that CRF19 most likely recombined around the 1970s in the Cuban community that was stationed in Democratic Republic of the Congo and Angola at that time. It was introduced very early into the Cuban province of Villa Clara, from where it had several introductions to La Habana in the 1980s and then further spread to other Cuban provinces. The CRF19 epidemic most probably started with heterosexual transmissions, followed in the 1980s by multiple introductions into "men having sex with men" (MSM) community, followed by multiple transmissions back to heterosexuals (often females). The early introduction of CRF19 into Cuba most likely explains its success, not observed in other parts of the world. However, importantly, its rapid progression to AIDS makes it crucial to survey CRF19 sub-epidemics not only in Cuba, but also in other parts of the world having regular exchanges with Cuba.

bioinformatics↗

bioRxiv2021

Analysis of HIV Reservoirs in Cellular Conjugates from Peripheral Blood

Defining distinctive attributes of HIV-infected cells will inform development of HIV cure-directed therapies. Prior ex vivo studies of blood and tissue have suggested that some HIV-infected CD4 T cells are found in conjugates with other cell types. Here, we analyzed levels and sequences of HIV nucleic acids in sorted cellular conjugates from PBMC. Compared to single CD4 T cells, conjugates containing CD4 T cells showed no enrichment for HIV DNA or RNA. However, in several HIV controllers, HIV DNA sequences from sorted conjugates were enriched for sequences closely related to plasma viruses. In ART-treated people, although subgenomic HIV DNA sequences in sorted conjugates and single cells were genetically intermingled, intact proviruses were more frequent in whole blood cells than in magnetically-purified CD4 T cells. We conclude that some HIV-infected cells have attributes that predict preferential loss during sample processing, and that may also reflect vulnerability to therapeutic targeting in vivo.

immunology↗

bioRxiv2020

Droplet microfluidic sequencing of HIV genomes and integration sites

Sequencing individual HIV-proviruses and their adjacent cellular junctions can elucidate mechanisms of infected cell persistence in vivo. Here, we present a high throughput microfluidic method to sequence entire proviruses in their native integration site context. We used the method to analyze infected cells from people with HIV on suppressive antiretroviral therapy, demonstrating >90% capture and sequencing of paired proviral genomes and integration sites. This method should enable comprehensive genetic analysis of persistent HIV-infected cell reservoirs, providing important insights into the barriers to HIV cure.

genomics↗

bioRxiv2020

Dog Savior: Immediate Scent-Detection of SARS-COV-2 by Trained Dogs

Molecular tests for viral diagnostics are essential to confront the COVID-19 pandemic, but their production and distribution cannot satisfy the current high demand. Early identification of infected people and their contacts is the key to being able to isolate them and prevent the dissemination of the pathogen; unfortunately, most countries are unable to do this due to the lack of diagnostic tools. Dogs can identify, with a high rate of precision, unique odors of volatile organic compounds generated during an infection; as a result, dogs can diagnose infectious agents by smelling specimens and, sometimes, the body of an infected individual. We trained six dogs of three different breeds to detect SARS-CoV-2 in respiratory secretions of infected patients and evaluated their performance experimentally, comparing it against the gold standard (rRT-PCR). Here we show that viral detection takes one second per specimen. After scent-interrogating 9,200 samples, our six dogs achieved independently and as a group very high sensitivity, specificity, predictive values, accuracy, and likelihood ratio, with very narrow confidence intervals. The highest metric was the negative predictive value, indicating that with a disease prevalence of 7.6%, 99.9% of the specimens indicated as negative by the dogs did not carry the virus. These findings demonstrate that dogs could be useful to track viral infection in humans, allowing COVID-19 free people to return to work safely.

microbiology↗

bioRxiv2020

Expression and T cell Regulatory Action of the PD-1 Immune Checkpoint in the Ovary and Fallopian Tube

The Programmed Cell Death Protein-1 (PD-1/PDCD-1/CD279) checkpoint has powerful immunomodulatory action, including in the context of cancer. PD-1 receptor activation by its ligands (PD-L1/2) is associated with downregulated immune response, and tumor cells can avoid surveillance via PD-1 and/or ligand expression. While receptor expression is largely limited to lymphoid, myeloid, and tumor cells, we show that membrane bound and soluble variants of PD-1 and ligands are also expressed by permanent constituent cell types of the human ovary and fallopian tube, including granulosa cells and oocytes. PD-1 and soluble ligands were highly enriched in exosome fractions in human follicular fluid at bioactive levels that can control T cell PD-1 activation. PD-1 checkpoint signaling may be involved in physiological ovarian functions including follicle, and ultimately, germline and embryo immune-privilege.Competing Interest StatementThe authors have declared no competing interest.View Full Text

physiology↗

bioRxiv2020

In silico analysis of RT-qPCR designs recommended by WHO for detection of SARS-CoV-2 and a commercial kit validated following UNE/EN ISO 17025:2005 and two reference laboratories

BackgroundThe Corona Virus Disease 2019 (COVID-19), caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), has become a serious infectious disease affecting human health worldwide and rapidly declared a pandemic by WHO. Early, several RT-qPCR were designed by using only the first SARS-CoV-2 genome sequence. ObjectivesA few days later, when additional SARS-CoV-2 genome were retrieved, the kit GPS CoVID-19 dtec-RT-qPCR Test was designed to provide a highly specific detection method and commercially available worldwide. The kit was validated following criteria recommended by the UNE/EN ISO 17025:2005 and ISO/IEC 15189:2012. MethodsThe present study approached the in silico specificity of the GPS CoVID-19 dtec-RT-qPCR Test and RT-qPCR designs currently published. The empirical validation parameters specificity (inclusivity/exclusivity), quantitative phase analysis (10-106 copies), reliability (repeatability/reproducibility) and sensitivity (detection/quantification limits) were evaluated for a minimum of 10-15 assays. Diagnostic validation was achieved by two independent reference laboratories, the Instituto de Salud Carlos III (ISCIII), (Madrid, Spain) and the Public Health England (PHE; Colindale, London, UK). ResultsThe GPS RT-qPCR primers and probe showed the highest number of mismatches with the closet related non-SARS-CoV-2 coronavirus, including some indels. The kits passed all parameters of validation with strict acceptance criteria. Results from reference laboratories 100% correlated with these obtained by suing reference methods and received an evaluation with 100% of diagnostic sensitivity and specificity. ConclusionsThe GPS CoVID-19 dtec-RT-qPCR Test, available with full analytical and diagnostic validation, represents a case of efficient transfer of technology being successfully used since the pandemic was declared. The analysis suggested the GPS CoVID-19 dtec-RT-qPCR Test is the more exclusive by far.

genetics↗

bioRxiv2020

AncesTree: an interactive immunoglobulin lineage tree visualizer

High-throughput sequencing of human immunoglobulin genes allows analysis of antibody repertoires and the reconstruction of clonal lineage evolution. Phylip, an algorithm that has been originally developed for applications in ecology and macroevolution, can also be used for the phylogenic reconstruction of antibodies maturation pathway. The study of antibodies (Abs) affinity maturation is of specific interest to understand the generation of Abs with high affinity or broadly neutralizing activities. Phylogenic analysis enables the identification of the key somatic mutations required to achieve optimal antigen binding. To complement Phylip algorithm, we developed AncesTree, a graphic user interface (GUI) that aims to give researchers the opportunity to interactively explore antibodies clonal evolution. AncesTree displays interactive immunoglobulins (Ig) phylogenic tree, Ig related mutations and sequence alignments using additional information coming from specialized antibody tools (such as IMGT(R)). The GUI is a Java standalone application allowing interaction with Ig-tree that can run under Windows, Linux and Mac OS.

bioinformatics↗

bioRxiv2019

Malaria transmission assisted by interaction between Plasmodium α-tubulin-1 and Anopheles FREP1 protein

Passage of Plasmodium through a mosquito midgut is essential for malaria transmission. FREP1, a peritrophic matrix protein in a mosquito midgut, binds to the parasite and mediates Plasmodium infection in Anopheles. The FREP1-mediated Plasmodium invasion pathway is highly conserved across multiple species of Plasmodium and Anopheles. Through pulldown, nine P. berghei proteins were co-precipitated with FREP1-conjugated beads. After cloning these nine genes from P. berghei and expressing them in insect cells, six of them were confirmed to interact with recombinant FREP1 protein. Among them, -tubulin-1 and heat shock protein 70 (Hsp70) were highly conserved in Plasmodium species with >95% identity. Thus, P. falciparum -tubulin-1 and Hsp70 were cloned and expressed in E. coli to stimulate antibody (Ab) in mice. Our results showed that anti-serum against P. falciparum -tubulin-1 significantly inhibited P. falciparum transmission to An. gambiae, while Ab against P. falciparum Hsp70 serum did not. The polyclonal Ab against human -tubulin did not interfere formation of ookinetes, however, significantly reduced the number of P. falciparum oocysts in An. gambiae midguts. Moreover, fluorescence microscope assays showed that anti--tubulin Ab bound to impermeable Plasmodium ookinete apical invasive apparatus. Therefore, we propose that the interaction between Anopheles FREP1 protein and Plasmodium -tubulin-1 directs the ookinete invasive apparatus towards midgut peritrophic matrix for the efficient passage of the parasite. Anopheles FREP1 and Plasmodium -tubulin-1 are potential targets for blocking malaria transmission to the mosquito host. AUTHOR SUMMARYThe molecular mechanisms of malaria transmission to mosquito are not well-understood. FREP1 proteins in mosquito midget PM has been proved to mediate malaria transmission by binding to parasite ookinetes. Here we reported that Plasmodium parasite -tubulin-1 is an FREP1 binding partner. We initially identified the -tubulin-1 through the FREP1-pulldown assay; Then we cloned P. falciparum -tubulin-1, and demonstrated that the insect cell expressed recombinant Plasmodium -tubulin-1 bound to FREP1 in vitro; Next, mouse anti-serum against P. falciparum -tubulin-1 was found to inhibit P. falciparum transmission to An. gambiae. P. falciparum -tubulin-1 shares >84% identical amino acid sequence with human -tubulin, purified Ab against human -tubulin significantly inhibited malaria transmission. Anti-human -tubulin Ab did not interfere the gametocyte-to-ookinetes conversion. Final, we found that anti--tubulin Ab bound to the apical end of impermeable ookinetes. Structurally, ookinete invasive apparatus locates at the apical opening. Therefore, we propose that the interaction between Anopheles midgut FREP1 protein and Plasmodium apical -tubulin-1 directs the ookinete invasive apparatus towards midgut PM for the efficient parasite invasion.

microbiology↗

bioRxiv2018

Ask1 and Akt act synergistically to promote ROS-dependent regeneration in Drosophila

The mechanism by which apoptotic cells release signals that induce undamaged neighbor cells to proliferate and regenerate missing parts remains elusive. Oxidative stress originated by dying or damaged cells can be propagated to neighboring cells, which then promote regeneration. We investigated the nature of the stress sensing mechanism by which neighboring cells are recruited. We found that Drosophila apoptosis signal-regulating kinase 1 (Ask1) senses reactive oxygen species (ROS) differently in stressed dying cells and unstressed neighboring cells and this differential sensing is pivotal for tissue repair. In undamaged cells, this activity is attenuated, but not abolished, by Akt1 phosphorylation, which thus acts as a survival signal that results in the tolerable levels of p38 and JNK necessary for regeneration. These observations demonstrate that the non-autonomous activation of the ROS-sensing mechanism by Ask1 and Akt1 in neighboring unstressed cells. Collectively, these results provide the basis for understanding the molecular mechanism of communication between dying and living cells that triggers regeneration.\n\nAuthor summaryOne of the early events that occur after tissue damage is oxidative stress production that signals to initiate wound healing and regeneration. Several signaling pathways, such as JNK and p38, respond to oxidative stress and are necessary for regeneration. We decided to explore the mechanism that links the oxidative stress and the activation of these pathways. We used epithelia of Drosophila to genetically direct cell death in specific zones of the tissue as means of experimentally controlled cell damage. We found that the Ask1 protein, which is sensitive to oxidative stress, is a key player in this scenario. Actually it acts as an intracellular sensor that upon damage activates those signaling pathways. However, high activity of Ask1 can be toxic for the cell. This is controlled by Akt, an enzyme dowstream the insulin pathway, with attenuates the activity of Ask1 to tolerable levels. In conclusion, Ask1 and Akt act synergistically to respond to the stress generated after tissue damage and drive regeneration. In other words, we found that the link between oxidative stress and nutrition is key for tissue regeneration.

developmental biology↗

bioRxiv2017

Large scale gene duplication affected the European eel (Anguilla anguilla) after the 3R teleost duplication

Genomic scale duplication of genes generates raw genetic material, which may facilitate new adaptations for the organism. Previous studies on eels have reported specific gene duplications, however a species-specific large-scale gene duplication has never before been proposed. In this study, we have assembled a de novo European eel transcriptome and the data show more than a thousand gene duplications that happened, according to a 4dTv analysis, after the teleost specific 3R whole genome duplication (WGD). The European eel has a complex and peculiar life cycle, which involves extensive migration, drastic habitat changes and metamorphoses, all of which could have been facilitated by the genes derived from this large-scale gene duplication.\n\nOf the paralogs created, those with a lower genetic distance are mostly found in tandem repeats, indicating that they are young segmental duplications. The older eel paralogs showed a different pattern, with more extensive synteny suggesting that a Whole Genome Duplication (WGD) event may have happened in the eel lineage. Furthermore, an enrichment analysis of eel specific paralogs further revealed GO-terms typically enriched after a WGD. Thus, this study, to the best of our knowledge, is the first to present evidence indicating an Anguillidae family specific large-scale gene duplication, which may include a 4R WGD.

genomics↗