Neisseria gonorrhoeae subverts formin-dependent actin polymerization to colonize human macrophages

Dynamic reorganization of the actin cytoskeleton dictates plasma membrane morphogenesis and is frequently subverted by bacterial pathogens for entry and colonization of host cells. The human-adapted bacterial pathogen Neisseria gonorrhoeae can colonize and replicate when cultured with human macrophages, however the basic understanding of how this process occurs is incomplete. N. gonorrhoeae is the etiological agent of the sexually transmitted disease gonorrhea and tissue resident macrophages are present in the urogenital mucosa, which is colonized by the bacteria. We uncovered that when gonococci colonize macrophages, they can establish an intracellular or a cell surface-associated niche that support bacterial replication independently. Unlike other intracellular bacterial pathogens, which enter host cells as single bacterium, establish an intracellular niche and then replicate, gonococci invade human macrophages as a colony. Individual diplococci are rapidly phagocytosed by macrophages and transported to lysosomes for degradation. However, we found that surface-associated gonococcal colonies of various sizes can invade macrophages by triggering actin skeleton rearrangement resulting in plasma membrane invaginations that slowly engulf the colony. The resulting intracellular membrane-bound organelle supports robust bacterial replication. The gonococci-occupied vacuoles evaded fusion with the endosomal compartment and were enveloped by a network of actin filaments. We demonstrate that gonococcal colonies invade macrophages via a process mechanistically distinct from phagocytosis that is regulated by the actin nucleating factor FMNL3 and is independent of the Arp2/3 complex. Our work provides insights into the gonococci life-cycle in association with human macrophages and defines key host determinants for macrophage colonization.