Biology subjects

Frangieh, J.

Publications and source records attributed to Frangieh, J..

3 recordsLinked to original sources

bioRxiv2026

Covalently linked peptides and membrane potential enable CyaA segment translocation

The adenylate cyclase toxin (CyaA) from Bordetella pertussis intoxicates host cells by directly translocating its N-terminal catalytic domain across the plasma membrane; however, the forces driving this unique process remain poorly defined. Here, we dissect the membrane translocation mechanisms of two peptide segments derived from CyaA: P233 and P454 from the catalytic domain and the translocation region, respectively. Both P454 and P233 are calmodulin-binding segments that are sequentially involved in the translocation and activation of the catalytic domain. Using a newly developed Droplet Interface Bilayer (DIB) approach, called DIB-Pipette, which enables direct visualization of peptide transport under controlled membrane potentials, we show that P454 translocates across membranes independently of membrane potential, whereas P233 translocation requires a negative electric membrane potential. Strikingly, covalent coupling of P233 and P454 enables efficient translocation of the resulting peptide even in the absence of a membrane potential. Together, these results suggest that two distinct membrane-active segments within CyaA act cooperatively to promote translocation at the peptide level, revealing an intrinsic mechanism that may contribute to membrane potential-dependent translocation. These findings provide new mechanistic insights into CyaA cell intoxication process and reveal a multifunctional strategy for protein delivery across membranes. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=159 SRC="FIGDIR/small/716334v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@750ab3org.highwire.dtl.DTLVardef@11a980org.highwire.dtl.DTLVardef@18f2b27org.highwire.dtl.DTLVardef@5a3a59_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2025

Post-translational acylation drives folding and activity of the CyaA bacterial toxin.

Post-translational modifications critically shape protein conformation and function, yet how they regulate bacterial toxins remains elusive. The adenylate cyclase (CyaA) toxin is a major virulence factor of Bordetella pertussis, the causative agent of whooping cough. CyaA is produced as an inactive precursor, proCyaA, which is activated by acylation of two lysine residues within the bacterium. Once acylated and secreted, CyaA invades innate immune cells and disrupts their phagocytic functions. High-resolution structural characterization of CyaA has remained elusive due to its size, multi-domain organization, flexibility, and aggregation propensity. Here, we overcome these challenges and generate the first structural ensembles of both non-acylated and acylated CyaA in solution by combining experimental data with integrative modeling. Coarse-grained molecular dynamics simulations reveal that acylation is critical to stabilize the native fold and to favorably orient CyaA on the target membrane. Overall, our findings reveal how post-translational acylation triggers native folding and provide mechanistic insights into the early steps of host cell intoxication.

biophysics↗

bioRxiv2023

The interpeduncular nucleus blunts the rewarding effect of nicotine

Nicotine, by stimulating ventral tegmental area (VTA) dopaminergic neurons, has a rewarding effect that drives tobacco consumption. In turn, the interpeduncular nucleus (IPN) is thought to become activated at high nicotine doses to restrict drug intake. However, the dynamics of the IPN response to nicotine and its impact on the rewarding effect of the drug remain unknown. To address this issue, we have developed a genetically-modified mouse model, in which a "suicide" antagonist of nicotinic acetylcholine receptors (nAChRs) selectively attaches to a designer {beta}4 nAChR subunit. By locally infusing this antagonist in the IPN, we achieved pharmacologically-specific and sustained antagonism of nAChRs containing the {beta}4 subunit. By combining this chemogenetic method with in vivo electrophysiology, we show that even at low doses, nicotine activates and inhibits two different populations of IPN neurons, and that {beta}4-containing nAChRs are only involved in the activation response. Furthermore, blocking the response to nicotine selectively in the IPN increased both the sensitivity of the VTA to the drug and its rewarding effect in a conditioned place preference paradigm. These findings indicate that the IPN is engaged across a large range of nicotine doses and acts as a regulatory brake on the nicotine reward circuit.

neuroscience↗