Legionella pneumophila , the pathogen responsible for Legionnaires’ disease, relies on the Dot/Icm type IV secretion system (T4SS) to establish infection. This complex machine is composed of more than 30 proteins and delivers over 300 protein substrates into host cells to disrupt immune defenses and create a replicative niche. At the center of this process is the T4SS coupling complex, which mediates substrate recognition and transport. Within this complex, the IcmSW proteins play a critical role in recruiting effector proteins for delivery. In this study, these effector-IcmSW complexes will be purified and biophysically characterized to define the molecular details of substrate engagement. For the first time, purified effector-IcmSW complexes will be structurally characterized using single particle cryo-electron microscopy (cryo-EM) to reveal how effectors are engaged. Furthermore, we are investigating how ATPases promote the translocation of these effector proteins through the secretion system, and cryo-EM has been used to identify structural features that may provide new insight into the overall mechanism of effector transport. By reconstituting functional protein complexes under controlled conditions, this work will move beyond traditional cell-based assays and provide detailed insight into how the Dot/Icm system selects and delivers its diverse set of effectors. These findings will advance understanding of bacterial protein transport across membrane barriers and reveal fundamental principles of macromolecule translocation in pathogenic bacteria.
Somarathne et al. (Sun,) studied this question.