Biology subjects

Hyrien, O.

Publications and source records attributed to Hyrien, O..

16 recordsLinked to original sources

bioRxiv2026

A Multiclass Mechanistic Model of Vaccine-Induced Humoral Response

Vaccine-induced humoral immunity emerges from complex interactions among competing B-cell populations and antibody production in response to antigen exposure, yet the mechanisms governing these interactions remain only partially understood. We developed a mechanistic mathematical model that integrates vaccine regimen features, immunogen kinetics, multiclass B-cell dynamics, and germinal center processes. The model was calibrated to data from the G001 HIV vaccine trial evaluating eOD-GT8, a germline-targeting immunogen designed to activate VRC01-class precursor B cells capable of developing into broadly neutralizing antibody-producing cells. The model reproduced key features of the observed B-cell and antibody responses, enabled estimation of class-specific activation and competition parameters, and supported investigation of the identifiability of competition parameters. Activation rates varied across B-cell classes, with memory B cells exhibiting substantially higher activation rates than naive B cells. Importantly, competition was stronger within than between classes, with the strongest competition occurring among VRC01-class B cells. These findings suggest that class-specific differences in activation and competition shape the magnitude and composition of vaccine-induced humoral response and may influence the expansion of broadly neutralizing antibody precursors. More broadly, the model provides a quantitative framework for dissecting multiclass B-cell competition and antibody responses, and for evaluating how vaccination strategies shape humoral immunity.

immunology↗

bioRxiv2026

High-resolution single-molecule replication profiling of the human genome

Despite the considerable progress made in recent years thanks to the rise of long-read sequencing, high-resolution, single molecule (SM)-based replication profiling of large genomes has remained out of reach. Here, we present a replication labelling strategy consisting in repeated pulse-labelling of asynchronously growing cells with the thymidine analogue bromodeoxyuridine (BrdU) that greatly enhances the number of detectable replication tracks. We show that the multipulse BrdU labelling protocol, in association with ForkML, a machine-learning method translating BrdU signals in nanopore reads into oriented and positioned replication forks, enables the establishment of a SM replication map of the entire human genome at kilobase resolution.

genomics↗

bioRxiv2026

A Closed-Form Bayesian Framework for DNA Replication Reveals Intrinsic Origin Timing and Activation Delays

I.We present an analytical framework for modeling eukaryotic DNA replication that, given experimental Replication Fork Directionality (RFD) data, enables Bayesian inference of origin number, activation delay [Formula] and intrinsic timing ({lambda}i), (the mean replication time if each origin were isolated). By deriving closed-form expressions for RFD and Mean Replication Timing (MRT) under exponential and a specific Weibull firing-time distributions as functions of [Formula] and ({lambda}i), we eliminate the need for stochastic simulations. These analytical results reveal that RFD, as a ratio of fork directions, is invariant under joint rescaling of intrinsic timing and fork speed; absolute intrinsic timing can nonetheless be inferred when fork speed is independently measured. We demonstrate that under exponential firing distribution for the origin, the observed efficiency (Ei), i.e. the probability for an origin to fire which accounts for nearby origin, is simply MRT(xi)/{lambda}i. The closed-form RFD expressions allow to use a Bayesian method that achieves 0.96-0.99 correlation with yeast RFD profiles and resolves ~780 origins in S. cerevisiae. Our framework identifies about 150 origins with biologically significant delays ([≥] 3 minute), revealing regulated activation kinetics undetectable by existing methods. By quantifying how origin intrinsic timing and delays shape replication timing landscapes, this work confirms yeast as a paradigm organism for studying DNA replication control mechanisms.

biophysics↗

bioRxiv2025

Engineering Sustained-Release Broadly Neutralizing Antibody Formulations

Sustained serum levels of broadly neutralizing antibodies (bnAbs) are crucial for effective passive immunization against infectious diseases as protection persists only while these bnAbs remain at adequate concentrations within the body. Current obstacles, such as poor pharmacokinetics (PK) and burdensome administration, must be overcome to make bnAbs a viable option for pre- and post-exposure prophylaxis. In this work, we explore how a polymer-nanoparticle (PNP) hydrogel depot technology can be engineered to prolong protein delivery. In-vivo studies in mice and rats demonstrate prolonged protein release, and modeling efforts predict the impact of both the elimination half-life of the active pharmaceutical ingredient and hydrogel depot volume on overall pharmacokinetics. Moreover, flow cytometry characterization reveals that immune cell infiltration into the hydrogel depot can result in faster-than-expected release of antibody cargo on account of active transport via cellular uptake. We then demonstrate that co-formulation of antibodies with an anti-inflammatory agent reduces cellular infiltration and resulting active transport, further extending delivery and pharmacokinetics. Finally, multicompartmental modeling predicts the human PK profiles of clinically relevant HIV bnAbs delivered via subcutaneous hydrogel injection. These findings aid in the development of next generation hydrogel materials that stabilize and slowly release bnAbs for long-term pre-exposure immunoprophylaxis.

bioengineering↗

bioRxiv2024

High Resolution Class I HLA -A, -B, and -C Diversity in Eastern and Southern African Populations

Africa remains significantly underrepresented in high-resolution Human Leukocyte Antigen (HLA) data, despite being one of the most genetically diverse regions in the world. This critical gap in genetic information poses a substantial barrier to HLA-based research on the continent. In this study, Class I HLA data from Eastern and Southern African populations were analysed to assess genetic diversity across the region. We examined allele and haplotype frequency distributions, deviations from Hardy-Weinberg Equilibrium (HWE), linkage disequilibrium (LD), and conducted neutrality tests of homozygosity across various populations. Additionally, the African HLA data were compared to those of Caucasian and African American populations using the Jaccard index and multidimensional scaling (MDS) methods. The study revealed that South African populations exhibited 50.4% more genetic diversity within the Class I HLA region compared to other African populations. Zambia showed an estimated 36.5% genetic diversity, with Kenya, Rwanda and Uganda showing 35.7%, 34.2%, and 31.1%, respectively. Furthermore, an analysis of in-country diversity among diferent tribes indicated an average Class I HLA diversity of 25.7% in Kenya, 17% in Rwanda, 2.8% in South Africa, 13.6% in Uganda, and 6.5% in Zambia. The study also highlighted the genetic distinctness of Caucasian and African American populations compared to African populations. Notably, the diferential frequencies of disease- promoting and disease-preventing HLA alleles across these populations emphasize the urgent need to generate high-quality HLA data for all regions of Africa and its major ethnic groups. Such eforts will be crucial in enhancing healthcare outcomes across the continent. Author SummaryThis study investigated the diversity of class I HLA in the eastern and southern regions of the African continent using a population genetics approach. Analysis of HLA data at both country and tribal levels revealed significant genetic diferences and the unique characteristics of these populations compared to Caucasian and African American populations in the United States. The diferential frequencies of disease-promoting and disease-preventing HLA alleles across these populations suggest that large-scale vaccine administration may be inefective without a thorough understanding of the HLA composition of each population. This study highlights the urgent need to generate high-quality HLA data across all regions of Africa and its major ethnic groups. Such comprehensive data collection is essential for optimizing vaccine design, deepening our understanding of HLA-disease associations, and ultimately improving healthcare outcomes across the continent.

genetics↗

bioRxiv2024

Nanotiming: telomere-to-telomere DNA replication timing profiling by nanopore sequencing

Current temporal studies of DNA replication are either low-resolution or require complex cell synchronisation and/or sorting procedures. Here we introduce Nanotiming, a nanopore sequencing-based method producing high-resolution, telomere-to-telomere replication timing (RT) profiles of eukaryotic genomes by interrogating changes in intracellular dTTP concentration during S phase through competition with its analogue bromodeoxyuridine triphosphate (BrdUTP) for incorporation into replicating DNA. Nanotiming solely demands the labelling of asynchronously growing cells with an innocuous dose of BrdU during one doubling time followed by BrdU quantification along nanopore reads. We demonstrate in yeast S. cerevisiae that Nanotiming precisely reproduces RT profiles generated by reference methods in wild-type and mutant cells inactivated for known RT determinants, for one-tenth of the cost. Nanotiming is simple, accurate, inexpensive, amenable to large-scale analyses, and is capable of unveiling RT at individual telomeres, revealing that Rif1 iconic telomere regulator directly delays the replication only of telomeres with specific subtelomeric elements.

genomics↗

bioRxiv2024

Dual DNA Replication Modes: Varying Fork Speeds and Initiation Rates within the spatial replication program in Xenopus

Large vertebrate genomes duplicate by activating tens of thousands of DNA replication origins, irregularly spaced along the genome. The spatial and temporal regulation of the replication process is not yet fully understood. To investigate the DNA replication dynamics, we developed a methodology called RepliCorr, which uses the spatial correlation between replication patterns observed on stretched single-molecule DNA obtained by either DNA combing or high-throughput optical mapping. The analysis revealed two independent spatiotemporal processes that regulate the replication dynamics in the Xenopus model system. These mechanisms are referred to as a fast and a slow replication mode, differing by their opposite replication fork speed and rate of origin firing. We found that Polo-like kinase 1 (Plk1) depletion abolished the spatial separation of these two replication modes. In contrast, neither replication checkpoint inhibition nor Rif1 depletion affected the distribution of these replication patterns. These results suggest that Plk1 plays an essential role in the local coordination of the spatial replication program and the initiation-elongation coupling along the chromosomes in Xenopus, ensuring the timely completion of the S phase.

bioinformatics↗

bioRxiv2022

AAV-delivered gene editing for latent genital or orofacial herpes simplex virus infection reduces ganglionic viral load and minimizes subsequent viral shedding in mice

Herpes simplex virus (HSV) establishes latency in ganglionic neurons of the peripheral nervous system, from which it can reactivate, causing recurrent disease and possible transmission to a new host. Current anti-HSV therapy does not eliminate latent HSV, and thus is only suppressive rather than curative. We developed a potentially curative approach to latent HSV infection and pathogenesis, based on gene editing using HSV-specific meganucleases delivered by adeno-associated virus (AAV) vectors. Our results demonstrated that a dual meganuclease therapy, composed of two anti-HSV-1 meganucleases delivered by a triple AAV serotype combination (AAV9, AAV-Dj/8, AAV-Rh10), can eliminate up to 97% of latent HSV DNA from ganglia in both ocular and vaginal mouse models of latent HSV infection. Using a novel pharmacological approach to reactivate latent HSV-1 in mice with the bromodomain inhibitor JQ-1, we demonstrated that this reduction in ganglionic viral load leads to a significant reduction of viral shedding from treated vs. control mice, with many treated mice showing no detectable virus shedding. In general, therapy was well tolerated, although dose-ranging studies showed hepatotoxicity at high AAV doses, consistent with previous observations in animals and humans. Also in agreement with previous literature, we observed subtle histological evidence of neuronal injury in some experimental mice, although none of the mice demonstrated observable neurological signs or deficits. These results reinforce the curative potential of gene editing for latent orofacial and genital HSV disease, and provide a framework for additional safety studies before human trials can begin.

neuroscience↗

bioRxiv2021

Inferring the DNA replication origin density landscape in human cells from replication timing and fork directionality data

In human and other metazoans, the determinants of replication origin location and strength are still elusive. Origins are licensed in G1 phase and fired in S phase of the cell cycle, respectively. It is debated which of these two temporally separate steps determines origin efficiency. Experiments can independently profile mean replication timing (MRT) and replication fork directionality (RFD) genome-wide. Such profiles contain information on multiple origins properties and on fork speed. Due to possible origin inactivation by passive replication, however, observed and intrinsic origin efficiencies can markedly differ. Thus, there is a need for methods to infer intrinsic from observed origin efficiency, which is context-dependent. Here, we show that MRT and RFD data are highly consistent with each other but contain information at different spatial scales. Using neural networks, we infer an origin licensing landscape that, when inserted in an appropriate simulation framework, jointly predicts MRT and RFD data with unprecedented precision and underlies the importance of dispersive origin firing. We furthermore uncover an analytical formula that predicts intrinsic from observed origin efficiency combined with MRT data. Comparison of inferred intrinsic origin efficiencies with experimental profiles of licensed origins (ORC, MCM) and actual initiation events (Bubble-seq, SNS-seq, OK-seq, ORM) show that intrinsic origin efficiency is not solely determined by licensing efficiency. Thus, human replication origin efficiency is set at both the origin licensing and firing steps. Author SummaryDNA replication is a vital process that produces two identical replicas of DNA from one DNA molecule, ensuring the faithful transmission of genetic information from mother to daughter cells. The synthesis of new DNA strands initiates at multiple sites, termed replication origins, propagates bidirectionally, and terminates by merging of converging strands. Replication initiation continues in unreplicated DNA but is blocked in replicated DNA. Experiments have only given partial information about origin usage. In this work we reveal the exact propensity of any site to initiate replication along human chromosomes. First, we simulate the DNA replication process using approximate origin information, predict the direction and time of replication at each point of the genome, and train a neural network to precisely recover from the predictions the starting origin information. Second, we apply this network to real replication time and direction data, extracting the replication initiation propensity landscape that exactly predicts them. We compare this landscape to independent origin usage data, benchmarking them, and to landscapes of protein factors that mark potential origins. We find that the local abundance of such factors is insufficient to predict replication initiation and we infer to which extent other chromosomal cues locally influence potential origin usage.

biophysics↗

bioRxiv2020

Human ORC/MCM density is low in active genes and correlates with replication time but does not solely define replication initiation zones

Eukaryotic DNA replication initiates during S phase from origins that have been licensed in the preceding G1 phase. Here, we compare ChIP-seq profiles of the licensing factors Orc2, Orc3, Mcm3, and Mcm7 with gene expression, replication timing and fork directionality profiles obtained by RNA-seq, Repli-seq and OK-seq. ORC and MCM are strongly and homogeneously depleted from transcribed genes, enriched at gene promoters, and more abundant in early-than in late-replicating domains. Surprisingly, after controlling these variables, no difference in ORC/MCM density is detected between initiation zones, termination zones, unidirectionally replicating and randomly replicating regions. Therefore, ORC/MCM density correlates with replication timing but does not solely regulate the probability of replication initiation. Interestingly, H4K20me3, a histone modification proposed to facilitate late origin licensing, was enriched in late replicating initiation zones and gene deserts of stochastic replication fork direction. We discuss potential mechanisms that specify when and where replication initiates in human cells.

cell biology↗

bioRxiv2020

Organization of DNA replication origin firing in Xenopus egg extracts : the role of intra-S checkpoint

During cell division, the duplication of the genome starts at multiple positions called replication origins. Origin firing requires the interaction of rate-limiting factors with potential origins during the S(ynthesis)-phase of the cell cycle. Origins fire as synchronous clusters which is proposed to be regulated by the intra-S checkpoint. By modelling the unchallenged, the checkpoint-inhibited and the checkpoint protein Chk1 over-expressed replication pattern of single DNA molecules from Xenopus sperm chromatin replicated in egg extracts, we demonstrate that the quantitative modelling of data requires: 1) a segmentation of the genome into regions of low and high probability of origin firing; 2) that regions with high probability of origin firing escape intra-S checkpoint regulation and 3) the variability of the rate of DNA synthesis close to replication forks is a necessary ingredient that should be taken in to account in order to describe the dynamic of replication origin firing. This model implies that the observed origin clustering emerges from the apparent synchrony of origin firing in regions with high probability of origin firing and challenge the assumption that the intra-S checkpoint is the main regulator of origin clustering. Author summaryDNA replication is one of the fundamental cell functions. The genome of eukaryotic organisms is duplicated from multiple positions named replication origins. Single molecule experiments allow to visualise the dynamics of spatio-temporal patterns created during replication process. The dynamic of replication process is regulated by checkpoints. However, the influence and the role of checkpoint regulation in the dynamics of spatio-temporal patterns of DNA replication is not understood. In this work we build a minimal, process-based and data rooted numerical model that allows to decipher the impact of checkpoint regulation on the dynamics of spatio-temporal pattern of DNA replication.

cell biology↗

bioRxiv2020

FORK-seq: replication landscape of the Saccharomyces cerevisiae genome by nanopore sequencing

Genome replication mapping methods profile cell populations, masking cell-to-cell heterogeneity. Here, we describe FORK-seq, a nanopore sequencing method to map replication of single DNA molecules at 200 nucleotide resolution. By quantifying BrdU incorporation along pulse-chased replication intermediates from Saccharomyces cerevisiae, we orient 58,651 replication tracks reproducing population-based replication directionality profiles and map 4,964 and 4,485 individual initiation and termination events, respectively. Although most events cluster at known origins and fork merging zones, 9% and 18% of initiation and termination events, respectively, occur at many locations previously missed. Thus, FORK-seq reveals the full extent of cell-to-cell heterogeneity in DNA replication.

genomics↗

bioRxiv2019

Active transcription regulates ORC/MCM distribution whereas replication timing correlates with ORC density in human cells

Eukaryotic replication initiates during S phase from origins that have been licensed in the preceding G1 phase. Here, we compare ChIP-seq profiles of the licensing factors Orc2, Orc3, Mcm3, and Mcm7 with replication initiation events obtained by Okazaki fragment sequencing. We demonstrate that MCM is displaced from early replicating, actively transcribed gene bodies, while ORC is mainly enriched at active TSS. Late replicating, H4K20me3 containing initiation zones display enhanced ORC and MCM levels. Furthermore, we find early RTDs being primarily enriched in ORC, compared to MCM, indicating that ORC levels are involved in organizing the temporal order of DNA replication. The organizational connection between active transcription and replication competence directly links changes in the transcriptional program to flexible replication patterns, which ensures the cells flexibility to respond to environmental cues.

cell biology↗

bioRxiv2018

Mapping DNA replication with nanopore sequencing

We have harnessed nanopore sequencing to study DNA replication genome-wide at the single-molecule level. Using in vitro prepared DNA substrates, we characterized the effect of bromodeoxyuridine (BrdU) substitution for thymidine on the MinION nanopore electrical signal. Using a neural-network basecaller trained on yeast DNA containing either BrdU or thymidine, we identified BrdU-labelled tracts in yeast cells synchronously entering S phase in the presence of hydroxyurea and BrdU. As expected, the BrdU-labelled tracts coincided with previously identified early-firing, but not late-firing, replication origins. These results open the way to high-throughput, high-resolution, single-molecule analysis of DNA replication in many experimental systems.

genomics↗

bioRxiv2018

Universal temporal rate of DNA replication origin firing: A balance between origin activation and passivation

The time-dependent rate I(t) of origin firing per length of unreplicated DNA presents a universal bell shape in eukaryotes that has been interpreted as the result of a complex time-evolving interaction between origins and limiting firing factors. Here we show that a normal diffusion of replication fork components towards localized potential replication origins (p-oris) can more simply account for the I(t) universal bell shape, as a consequence of a competition between the origin firing time and the time needed to replicate DNA separating two neighboring p-oris. We predict the I(t) maximal value to be the product of the replication fork speed with the squared p-ori density. We show that this relation is robustly observed in simulations and in experimental data for several eukaryotes. Our work underlines that fork-component recycling and potential origins localization are sufficient spatial ingredients to explain the universality of DNA replication kinetics.

biophysics↗

bioRxiv2017

High-throughput optical mapping of replicating DNA

DNA replication is a crucial process for the universal ability of living organisms to reproduce. Existing methods to map replication genome-wide use large cell populations and therefore smooth out variability between chromosomal copies. Single-molecule methods may in principle reveal this variability. However, current methods remain refractory to automated molecule detection and measurements. Their low throughput has therefore precluded genome-wide analyses. Here, we have repurposed a commercial optical DNA mapping device, the Bionano Genomics Irys system, to map the replication signal of single DNA molecules onto genomic position at high throughput. Our methodology (HOMARD) combines fluorescent labelling of replication tracks and nicking endonuclease (NE) sites with DNA linearization in nanochannel arrays and dedicated image processing. We demonstrate the robustness of our approach by providing an ultra-high coverage (23,311 x) replication map of bacteriophage {lambda} DNA in Xenopus egg extracts. HOMARD opens the way to genome-wide analysis of DNA replication at the single-molecule level.

genomics↗