Biology subjects

Villeneuve, J.

Publications and source records attributed to Villeneuve, J..

3 recordsLinked to original sources

bioRxiv2026

A novel IL-1β reporter assay identifies tolcapone as a caspase-1 suppressor targeting pyroptosis in endotoxemia

Unconventional protein secretion mediated by gasdermin D (GSDMD) pores is essential for the release of pro-inflammatory cytokines such as interleukin-1{beta} (IL-1{beta}), a major driver of many inflammatory pathologies. Despite extensive investigation over the years, discovery of IL-1{beta} secretion modulators has been hindered by the lack of robust, scalable experimental platforms. To date, IL-1{beta} related studies largely rely on primary cells and animal models, suitable for mechanistic studies but not readily scalable for high-throughput applications. Here, we engineered a CRISPR-based reporter cell line that allows quantitative monitoring of endogenous IL-1{beta} secretion while preserving the physiologically relevant inflammasome signaling. This platform faithfully recapitulated the response of primary macrophages to pathogen-associated molecular pattern (PAMPs) stimulation and supported the screening of an FDA-approved drug library comprising 1,398 compounds. Form this screen, we identified tolcapone as a potent inhibitor of IL-1{beta} secretion, reducing cytokine release by more than 80% across the screening pipeline. Mechanistically, tolcapone suppressed caspase-1 activation, thereby limiting GSDMD cleavage, pore formation, and the downstream maturation and secretion of IL-1{beta} and IL-18 in vitro. In vivo, tolcapone administration attenuated the acute inflammatory response in a lipopolysaccharide-induced endotoxemia model. Together, these findings establish our reporter platform as a robust tool for discovery of endogenous IL-1{beta} secretion modulators and identify tolcapone as a promising inhibitor of inflammasome-driven immune pathology.

immunology↗

bioRxiv2019

New factors for protein transport identified by a genome-wide CRISPRi screen in mammalian cells

Protein and membrane trafficking pathways are critical for cell and tissue homeostasis. Traditional genetic and biochemical approaches have shed light on basic principles underlying these processes. However, the list of factors required for secretory pathways function remains incomplete, and mechanisms involved in their adaptation poorly understood. Here, we present a powerful strategy based on a pooled genome-wide CRISPRi screen that allowed the identification of new factors involved in protein transport. Two newly identified factors, TTC17 and CCDC157, localized along the secretory pathway and were found to interact with resident proteins of ER-Golgi membranes. In addition, we uncovered that upon TTC17 knockdown, the polarized organization of Golgi cisternae was altered, creating glycosylation defects, and that CCDC157 is an important factor for the fusion of transport carriers to the Golgi complex. In conclusion, our work identified and characterized new actors in the mechanisms of protein transport and secretion, and opens stimulating perspectives for the use of our platform in physiological and pathological contexts.

cell biology↗

bioRxiv2019

Genome-wide CRISPR screening identifies new regulators of glycoprotein secretion

The fundamental process of protein secretion from eukaryotic cells has been well described for many years, yet gaps in our understanding of how this process is regulated remain. With the aim of identifying novel genes involved in the secretion of glycoproteins, we used a screening pipeline consisting of a pooled genome-wide CRISPR screen followed by secondary siRNA screening of the hits to identify and validate several novel regulators of protein secretion. We present approximately 50 novel genes not previously associated with protein secretion, many of which also had an effect on the structure of the Golgi apparatus. We further studied a small selection of hits to investigate their subcellular localisation. One of these, GPR161, is a novel Golgi-resident protein that we propose maintains Golgi structure via an interaction with golgin A5.

cell biology↗