Charlène Mireille Raymonde Raclot

De novo design of protein interactions with learned surface fingerprints

Michael Bronstein, Stephen M Buckley, Bruno Emanuel Ferreira De Sousa Correia, Beat Fierz, Pablo Gainza Cirauqui, Sandrine Madeleine Suzanne Georgeon, Casper Alexander Goverde, Zander Harteveld, Anthony Marchand, Jane Marsden, Dongchun Ni, Elisa Oricchio, Aaron Simone Petruzzella, Charlène Mireille Raymonde Raclot, Stéphane Rosset, Andreas Scheck, Henning Paul-Julius Stahlberg, Freyr Sverrisson, Alexandra Teslenko, Didier Trono, Priscilla Turelli, Alexandra Krina Van Hall-Beauvais, Sarah Wehrle

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 increase. This knowledge gap has been a major obstacle for the comprehensive understanding of cellular protein–protein interaction networks and for the de novo design of protein binders that are crucial for synthetic biology and translational applications. Here we use a geometric deep-learning framework operating on protein surfaces that generates fingerprints to describe geometric and chemical features that are critical to drive protein–protein interactions10. We hypothesized that 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, whereas others were generated purely 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 an approach for the de novo design of protein interactions and, more broadly, of artificial proteins with function.

2023

A high-throughput cell- and virus-free assay shows reduced neutralization of SARS-CoV-2 variants by COVID-19 convalescent plasma

Valeria Cagno, Jérémy Campos, Céline Pellaton, Florence Pojer, Charlène Mireille Raymonde Raclot, Didier Trono, Priscilla Turelli

The detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific antibodies in the serum of an individual indicates previous infection or vaccination. However, it provides limited insight into the protective nature of this immune response. Neutralizing antibodies recognizing the viral spike protein are more revealing, yet their measurement traditionally requires virus- and cell-based systems that are costly, time-consuming, inflexible, and potentially biohazardous. Here, we present a cell-free quantitative neutralization assay based on the competitive inhibition of trimeric SARS-CoV-2 spike protein binding to the angiotensin-converting enzyme 2 (ACE2) receptor. This high-throughput method matches the performance of the gold standard live virus infection assay, as verified with a panel of 206 seropositive donors with varying degrees of infection severity and virus-specific immunoglobulin G titers, achieving 96.7% sensitivity and 100% specificity. Furthermore, it allows for the parallel assessment of neutralizing activities against multiple SARS-CoV-2 spike protein variants of concern. We used our assay to profile serum samples from 59 patients hospitalized with coronavirus disease 2019 (COVID-19). We found that although most sera had high activity against the 2019-nCoV parental spike protein and, to a lesser extent, the alpha (B.1.1.7) variant, only 58% of serum samples could efficiently neutralize a spike protein derivative containing mutations present in the beta (B.1.351) variant. Thus, we have developed an assay that can evaluate effective neutralizing antibody responses to SARS-CoV-2 spike protein variants of concern after natural infection and that can be applied to characterize vaccine-induced antibody responses or to assess the potency of monoclonal antibodies.

AMER ASSOC ADVANCEMENT SCIENCE2021

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

Alexandre Coudray, Bart Deplancke, Julien Léonard Duc, Delphine Yvette L Grun, Evgeniia Pankevich, Christopher James Playfoot, Julien Paul André Pontis, Charlène Mireille Raymonde Raclot, Christian Axel Wandall Thorball, Didier Trono, Priscilla Turelli

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 co-opted 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 (ESCs). 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.

2020

A Dissection of Oligomerization by the TRIM28 Tripartite Motif and the Interaction with Members of the Krab-ZFP Family

Charlène Mireille Raymonde Raclot, Didier Trono, Priscilla Turelli

TRIM28 (also known as KAP1 or TIF1β) is the universal co-repressor of the Krüppel-associated box-containing zinc finger proteins (Krab-ZFPs), the largest family of transcription factors in mammals. During early embryogenesis, TRIM28 mediates the transcriptional silencing of many endogenous retroviral elements and genomic imprinted sites. Silencing is initiated by the recruitment of TRIM28 to a target locus by members of the Krab-ZFP. Subsequently, TRIM28 functions as a scaffold protein to recruit chromatin modifying effectors featuring SETDB1, HP1 and the NuRD complex. Although many protein partners involved in silencing have been identified, the molecular basis of the protein interactions that mediate silencing remains largely unclear. In the present study, we identified the first Bbox domain (T28_B1 135-203) as a molecular interface responsible for the formation of higher-order oligomers of TRIM28. The structure of this domain reveals a new interface on the surface of the Bbox domain. Mutants disrupting the interface disrupt the formation of oligomers but have no observed effect on transcriptional silencing defining a single TRIM28 dimer as the functional unit for silencing. Using assembly-deficient mutants, we employed small-angle X-ray scattering and biophysical techniques to characterize binding to member of the Krab-ZFP family. This allows us to narrow and define the binding interface to the center of the coiled-coil region (residues 294-321) of TRIM28 and define mutants that abolish binding to the Krab-ZFP proteins.

2019

Interplay of TRIM28 and DNA methylation in controlling human endogenous retroelements

Julien Léonard Duc, Adamantia Kapopoulou, Flavia Marzetta, Charlène Mireille Raymonde Raclot, Didier Trono, Priscilla Turelli

Reverse transcription-derived sequences account for at least half of the human genome. Although these retroelements are formidable motors of evolution, they can occasionally cause disease, and accordingly are inactivated during early embryogenesis through epigenetic mechanisms. In the mouse, at least for endogenous retroviruses, important mediators of this process are the tetrapod-specific KRAB-containing zinc finger proteins (KRAB-ZFPs) and their cofactor TRIM28. The present study demonstrates that KRAB/TRIM28-mediated regulation is responsible for controlling a very broad range of human-specific endogenous retroelements (EREs) in human embryonic stem (ES) cells and that it exerts, as a consequence, a marked effect on the transcriptional dynamics of these cells. It further reveals reciprocal dependence between TRIM28 recruitment at specific families of EREs and DNA methylation. It finally points to the importance of persistent TRIM28-mediated control of ERE transcriptional impact beyond their presumed inactivation by DNA methylation.

Cold Spring Harbor Lab Press, Publications Dept2014

The KRAB-ZFP/KAP1 System Contributes to the Early Embryonic Establishment of Site-Specific DNA Methylation Patterns Maintained during Development

Karolina Bojkowska, Adamantia Kapopoulou, Sandra Eloise Offner, Charlène Mireille Raymonde Raclot, Didier Trono, Priscilla Turelli

De novo DNA methylation is an essential aspect of the epigenetic reprogramming that takes place during early development, yet factors responsible for its instatement at particular genomic loci are poorly defined. Here, we demonstrate that the KRAB-ZFP-mediated recruitment of KAP1 to DNA in embryonic stem cells (ESCs) induces cytosine methylation. This process is preceded by H3K9 trimethylation, and genome-wide analyses reveal that it spreads over short distances from KAP1-binding sites so as to involve nearby CpG islands. In sharp contrast, in differentiated cells, KRAB/KAP1-induced heterochromatin formation does not lead to DNA methylation. Correspondingly, the methylation status of CpG islands in the adult mouse liver correlates with their proximity to KAP1-binding sites in ESCs, not in hepatocytes. Therefore, KRAB-ZFPs and their cofactor KAP1 are in part responsible for the establishment during early embryogenesis of site-specific DNA methylation patterns that are maintained through development.

Elsevier2012

A high-throughput cell- and virus-free assay shows reduced neutralization of SARS-CoV-2 variants by COVID-19 convalescent plasma

Valeria Cagno, Jérémy Campos, Céline Pellaton, Florence Pojer, Charlène Mireille Raymonde Raclot, Didier Trono, Priscilla Turelli

AMER ASSOC ADVANCEMENT SCIENCE2021

Charlène Mireille Raymonde Raclot

De novo design of protein interactions with learned surface fingerprints

Seroprevalence of anti-SARS-CoV-2 antibodies and cross-variant neutralization capacity after the Omicron BA.2 wave in Geneva, Switzerland: a population-based study

A high-throughput cell- and virus-free assay shows reduced neutralization of SARS-CoV-2 variants by COVID-19 convalescent plasma

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

A Dissection of Oligomerization by the TRIM28 Tripartite Motif and the Interaction with Members of the Krab-ZFP Family

Interplay of TRIM28 and DNA methylation in controlling human endogenous retroelements

The KRAB-ZFP/KAP1 System Contributes to the Early Embryonic Establishment of Site-Specific DNA Methylation Patterns Maintained during Development

De novo design of protein interactions with learned surface fingerprints

Seroprevalence of anti-SARS-CoV-2 antibodies and cross-variant neutralization capacity after the Omicron BA.2 wave in Geneva, Switzerland: a population-based study

Interplay of TRIM28 and DNA methylation in controlling human endogenous retroelements

A high-throughput cell- and virus-free assay shows reduced neutralization of SARS-CoV-2 variants by COVID-19 convalescent plasma

A Dissection of Oligomerization by the TRIM28 Tripartite Motif and the Interaction with Members of the Krab-ZFP Family

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

The KRAB-ZFP/KAP1 System Contributes to the Early Embryonic Establishment of Site-Specific DNA Methylation Patterns Maintained during Development