Biology subjects

Raclot, C.

Publications and source records attributed to Raclot, C..

12 recordsLinked to original sources

bioRxiv2026

An overexpression platform reveals the functional diversity of human KRAB-Zinc Finger Proteins in maintaining cellular homeostasis

Kruppel associated box zinc finger proteins (KZFPs) form the largest family of transcriptional regulators in mammals, yet most remain uncharacterized. Here we established a scalable framework to probe KZFP function. An arrayed inducible overexpression screen of 366 human KZFPs in K562 cells identified factors that alter cellular proliferation, enabling functional prioritization. Integrative transcriptomic, chromatin and proteomic analyses revealed diverse mechanisms, including transposable element-linked repression (ZNF43), promoter proximal regulation (ZNF257), and SCAN domain dependent transcriptional activation (ZNF498/ZSCAN25 and ZNF18). These results highlight the functional diversity of KZFPs and provide a strategy for their annotation.

genetics↗

bioRxiv2026

Antigenic landscape of a highly mutated SARS-CoV-2 Spike in ongoing viral evolution

The ongoing emergence of SARS-CoV-2 variants in an increasingly immune-experienced population is largely enabled by the plasticity of the Spike protein, which lies at the frontline of host immune pressure. Here, we investigated how extensive remodeling of the N-terminal domain (NTD) in Spike influences its antigenic properties. Using BA.2.87.1, a variant heavily mutated in this domain, we found that even large deletions do not substantially disrupt overall Spike structure or pseudovirus infectivity. However, our structural and binding analyses revealed that the NTD exhibits increased flexibility and that interaction with a heme-metabolite, contributing to assembly of an immunodominant epitope, is lost. These conformational effectscoupled with mutations, compromised the ability of human convalescent antibodies to engage this domain, contributing to their reduced neutralizing capacity. Consequently, BA.2.87.1-like variants may escape recognition by the pre-existing NTD-targeting antibodies, potentially reducing protection. Together, our results highlight the intrinsic adaptability of the Spike beyond the receptor-binding domain, with important implications for immune escape during viral evolution. TeaserStructural flexibility of the Spike and immune escape of a SARS-CoV-2 variant with public health implications.

immunology↗

bioRxiv2025

Cancer cells subvert the primate-specific KRAB zinc finger protein ZNF93 to control APOBEC3B

ZNF93 is a primate-restricted KRAB zinc finger protein responsible for repressing 20- to 12-million-year-old L1 transposable elements. Here, we reveal that ZNF93 also regulates the key cancer driver APOBEC3B--a mutagenic enzyme linked to tumorigenesis and cancer progression. ZNF93 depletion impairs DNA synthesis, activates replication and DNA damage checkpoints, and triggers proinflammatory phenotypes. Conversely, its overexpression enhances resistance to exogenous genotoxic stress, mirroring the effects observed with APOBEC3B depletion. ZNF93 expression correlates with cell proliferation rates and is overexpressed in many cancer types. These findings suggest that ZNF93 serves as a critical guardian of genome integrity, co-opted by cancer cells to counterbalance APOBEC3B-induced and L1-derived genomic instability and inflammation.

genetics↗

bioRxiv2025

Transposable element co-option drives transcription factor neofunctionalization

Gene duplication and domain acquisition are key mechanisms driving protein diversification. However, the impact of these processes on the function and evolution of transcription factors (TFs) remains poorly characterized. Here, we show that the DUF3669 domain, found in a subset of KRAB zinc-finger proteins (KZFPs), originated from the co-option of a fragment of a LINE-1 transposable element ORF1p. Similar to its LINE-1-encoded ancestor, the DUF3669 domain promotes KZFPs trimerization and enables the formation of nuclear condensates with ribonucleoparticle properties, which critically influence the genomic recruitment and action of these TFs. Thus, our study uncovers a direct link between TE co-option and TF neofunctionalization, highlighting how mobile genetic elements shape the evolution of protein functionality.

genetics↗

bioRxiv2025

Zinc-finger proteins with a co-opted capsid domain anchor nucleosomes over transposon sequences

Transposable elements (TEs) are frequently co-opted as cis-regulatory sequences that govern multiple aspects of host biology. The regulatory activity of these domesticated sequences are controlled by host factors, notably KRAB domain-containing zinc-finger proteins (KZFPs) in tetrapods. Here, we report that SCAN domain-containing zinc-finger proteins (SZFPs), which originally arose through capture of a retroviral capsid domain by a KZFP gene, have expanded and diversified their DNA recognition specificity to bind distinct TE subfamilies. We further demonstrate that SZFPs anchor nucleosomes at their target sites, and that their depletion leads to global shifts of nucleosomes away from underlying TE-derived sequences, occasionally accompanied by a gain of enhancer-associated chromatin states. Thus, SZFPs represent a novel layer of chromatin regulation centered on rapidly evolving TE-derived regulatory sequences.

genetics↗

bioRxiv2024

Evolutionarily recent transcription factors partake in human cell cycle regulation

The cell cycle is a fundamental process in eukaryotic biology and is accordingly controlled by a highly conserved core signaling cascade. However, whether recently evolved proteins also influence this process is unclear. Here, we systematically map the influence of evolutionarily recent transcription factors (TFs) on human cell cycle progression. We find that the genomic targets of select young TFs, many of which belong to the rapidly evolving Kruppel-associated box (KRAB) zinc-finger proteins (KZFP) family, exhibit synchronized cell cycle expression. Systematic perturbation studies reveal that silencing recent TFs disrupts normal cell cycle progression, which we experimentally confirm for ZNF519, a simian-restricted KZFP. Further, we show that the therian-specific KZFP ZNF274 sets the cell cycle expression and replication timing of hundreds of clustered genes. These findings highlight an underappreciated level of lineage specificity in cell cycle regulation.

cell biology↗

bioRxiv2022

P2G3 human monoclonal antibody neutralizes SARS-CoV-2 Omicron subvariants including BA.4 and BA.5 and Bebtelovimab escape mutants

The rapid evolution of SARS-CoV-2 has led to a severe attrition of the pool of monoclonal antibodies still available for COVID-19 prophylaxis or treatment. Omicron subvariants notably escape most antibodies developed so far, with Bebtelovimab last amongst clinically approved therapeutic antibodies to display still good activity against all of them including the currently dominant BA.4/BA.5. We recently described P2G3, a broadly active SARS-CoV-2 monoclonal antibody, which targets a region of Spike partly overlapping with the site recognized by Bebtelovimab. Here, we reveal that P2G3 efficiently neutralizes SARS-CoV-2 omicron subvariants including BA.4/BA.5. We further demonstrate that P2G3 neutralizes Omicron BA.2 and BA.4 mutants escaping Bebtelovimab blockade, whereas the converse is not true. FundingEU COVICIS program; private foundation advised by CARIGEST SA.

microbiology↗

bioRxiv2022

De novo design of site-specific protein interactions with learned surface fingerprints

Physical interactions between proteins are essential for most biological processes governing life. However, the molecular determinants of such interactions have been challenging to understand, even as genomic, proteomic, and structural data grows. This knowledge gap has been a major obstacle for the comprehensive understanding of cellular protein-protein interaction (PPI) networks and for the de novo design of protein binders that are crucial for synthetic biology and translational applications. We exploit a geometric deep learning framework operating on protein surfaces that generates fingerprints to describe geometric and chemical features critical to drive PPIs. We hypothesized these fingerprints capture the key aspects of molecular recognition that represent a new paradigm in the computational design of novel protein interactions. As a proof-of-principle, we computationally designed several de novo protein binders to engage four protein targets: SARS-CoV-2 spike, PD-1, PD-L1, and CTLA-4. Several designs were experimentally optimized while others were purely generated in silico, reaching nanomolar affinity with structural and mutational characterization showing highly accurate predictions. Overall, our surface-centric approach captures the physical and chemical determinants of molecular recognition, enabling a novel approach for the de novo design of protein interactions and, more broadly, of artificial proteins with function.

bioengineering↗

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

Structural analysis of the Spike of the Omicron SARS-COV-2 variant by Cryo-EM and implications for immune evasion

Investigation of potential hosts of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is crucial to understanding future risks of spillover and spillback. SARS-CoV-2 has been reported to be transmitted from humans to various animals after requiring relatively few mutations.[1] There is significant interest in describing how the virus interacts with mice as they are well adapted to human environments, are used widely as infection models and can be infected.[2] Structural and binding data of the mouse ACE2 receptor with the Spike protein of newly identified SARS-CoV-2 variants are needed to better understand the impact of immune system evading mutations present in variants of concern (VOC). Previous studies have developed mouse-adapted variants and identified residues critical for binding to heterologous ACE2 receptors.[3,4] Here we report the cryo-EM structures of mouse ACE2 bound to trimeric Spike ectodomains of four different VOC: Beta, Omicron BA.1, Omicron BA.2.12.1 and Omicron BA.4/5. These variants represent the oldest to the newest variants known to bind the mouse ACE2 receptor. Our high-resolution structural data complemented with bio-layer interferometry (BLI) binding assays reveal a requirement for a combination of mutations in the Spike protein that enable binding to the mouse ACE2 receptor. AUTHOR SUMMARYThe SARS-CoV-2 virus can infect different types of animals beyond humans. The virus uses its Spike protein on its surface to bind to cells. These cells have a protein called ACE2 that the Spike protein recognizes. Animals have slightly different ACE2 receptors compared to humans. Mice are widely used as a research animal and live in the same environments as humans so scientists are particularly interested. Understanding how Spike proteins binds to the mouse ACE2 receptor allows us to understand the impact of immune evading mutations found in new variants. We use a high resolution imaging technique called cryo-electron microscopy to look at how different Spike variants bind to the ACE2 receptor from mouse at a resolution where we can see the amino acids. We can see directly the individual amino acids and mutations on the Spike protein that interact with the mouse ACE2 receptor. Many of the mutations found in variants of concern also increase the strength of binding to the mouse ACE2 receptor. This result suggests that mutations in the Spike protein of future variants may have an additional effect in influencing how it binds to not only human ACE2 receptors but to mice and also different animals.

immunology↗

bioRxiv2021

S-acylation controls SARS-Cov-2 membrane lipid organization and enhances infectivity

SARS-CoV-2 virions are surrounded by a lipid bilayer which contains membrane proteins such as Spike, responsible for target-cell binding and virus fusion, the envelope protein E and the accessory protein Orf3a. Here, we show that during SARS-CoV-2 infection, all three proteins become lipid modified, through action of the S-acyltransferase ZDHHC20. Particularly striking is the rapid acylation of Spike on 10 cytosolic cysteines within the ER and Golgi. Using a combination of computational, lipidomics and biochemical approaches, we show that this massive lipidation controls Spike biogenesis and degradation, and drives the formation of localized ordered cholesterol and sphingolipid rich lipid nanodomains, in the early Golgi where viral budding occurs. ZDHHC20-mediated acylation allows the formation of viruses with enhanced fusion capacity and overall infectivity. Our study points towards S-acylating enzymes and lipid biosynthesis enzymes as novel therapeutic anti-viral targets.

cell biology↗

bioRxiv2019

Primate-restricted KRAB zinc finger proteins and target retrotransposons control gene expression in human neurons

In the first days of embryogenesis, transposable element-embedded regulatory sequences (TEeRS) are silenced by Kruppel-associated box (KRAB)-zinc finger proteins (KZFPs). Many TEeRS are subsequently coopted in transcription networks, but how KZFPs influence this process is largely unknown. We identify ZNF417 and ZNF587 as primate-specific KZFPs repressing HERVK (human endogenous retrovirus K) and SVA (SINE-VNTR-Alu) integrants in human embryonic stem cells (ESC). Expressed in specific regions of the human developing and adult brain, ZNF417/587 keep controlling TEeRS in ESC-derived neurons and brain organoids, secondarily influencing the differentiation and neurotransmission profile of neurons and preventing the induction of neurotoxic retroviral proteins and an interferon-like response. Thus, evolutionarily recent KZFPs and their TE targets partner up to influence human neuronal differentiation and physiology. One Sentence SummaryYoung transposable elements and their protein controllers team up to regulate the differentiation and function of human neurons.

evolutionary biology↗