Calcium sensor PefA times membrane repair to control mycobacterial vacuoles

Bacteria that live inside cells face an early challenge when they damage the membranes that surround them: the host cell must decide whether to patch the hole or send the damaged compartment for destruction. That decision is triggered by a universal early signal—calcium influx—but how calcium-sensing proteins coordinate the two major repair responses, ESCRT-mediated sealing and autophagy-based clearance, has been unclear. Sandra Guallar-Garrido and colleagues tackled this question using a tractable infection system, the Dictyostelium discoideum – Mycobacterium marinum infection model, which serves as a surrogate to study aspects of Mycobacterium tuberculosis intracellular pathogenesis. Combining genetic, imaging, and proteomic analyses, the team set out to find the calcium-responsive factors that guide the cell’s response when pathogen-containing vacuoles become damaged. Their work identified a previously unappreciated regulator: a penta-EF-hand protein named PefA, described as an ALG-2-like Ca 2+ sensor, that accumulates at damaged mycobacterial vacuoles and appears to play a central role in timing repair and autophagy decisions.

To trace how cells respond to vacuole damage, the researchers used the Dictyostelium discoideum – Mycobacterium marinum infection model and a suite of complementary approaches: genetic perturbations, live and fixed imaging, and proteomic analyses to follow protein recruitment. These methods revealed that PefA is transcriptionally upregulated during infection and concentrates at the mycobacterial vacuole. Functionally, PefA orchestrates recruitment of the E3 ubiquitin ligase TrafE, ESCRT components, and the autophagy machinery to damaged membranes. When PefA is present and active, vacuolar membranes are repaired in a timely manner, preserving vacuole integrity and supporting bacterial replication within the compartment. In contrast, loss of PefA impairs engagement of both ESCRT and autophagy pathways, leading to premature bacterial escape into the cytosol and altered infection outcomes. These results place PefA squarely at the intersection of damage sensing and the cellular repair toolkit.

The study uncovers a conserved Ca 2+ -dependent mechanism that links membrane damage sensing to the coordinated deployment of repair pathways. By identifying PefA, an ALG-2-like Ca 2+ sensor, as a timing device that recruits TrafE, ESCRT, and autophagy components, the work clarifies how cells balance sealing a damaged compartment against directing it to autophagy. That balance matters because it shapes host–pathogen interactions: whether a bacterium stays confined within a vacuole or escapes into the cytosol can change the course of infection. The findings have direct relevance to tuberculosis pathogenesis and host resilience to infection, highlighting a molecular node—PefA-regulated signaling—that could become the focus of further studies aiming to understand and possibly manipulate how infected cells respond to membrane damage.