Minute-scale control of ubiquitin-mediated degradation reveals dynamics of bacterial secreted effector-functions

Abstract

Precise temporal control of protein abundance is essential for dissecting dynamic cellular processes. While degron-based systems enable rapid protein depletion in eukaryotic cells, comparable tools are lacking for bacterial effectors delivered into host cells during infection. Here, we establish AIDE (Auxin-Inducible Degradation of Effectors), a host-directed degradation platform that harnesses the ubiquitin-proteasome system to selectively eliminate secreted bacterial proteins, including membrane-integrated effectors. By integrating a minimal auxin-inducible degron (AID) tag into effector genes, AIDE enables rapid, reversible, and spatially confined degradation while preserving native expression and secretion. Applied to Chlamydia trachomatis, AIDE revealed that the membrane-integrated deubiquitinase Cdu1 suppresses autophagy early and later promotes developmental transitions, whereas the integral membrane fusogen IncA is continuously required for inclusion integrity. This AIDE platform provides minute-scale, spatiotemporal control over bacterial effector activity, offering a broadly applicable framework for dissecting virulence mechanisms and host-pathogen interactions across diverse secretion-dependent pathogens.