ENKD1 attenuates antibacterial immunity by facilitating TRIM21-mediated RUBCN degradation to suppress LC3-associated phagocytosis

Microtubule-associated protein 1A/1B-light chain 3 (LC3)-associated phagocytosis (LAP) plays a critical role in host defense against invading pathogens, including <i><i>Listeria</i> monocytogenes (<i>Listeria. monocytogenes</i>)</i>, <i><i>Salmonella typhimurium (<i>S. typhimurium</i>)</i></i>, and <i>Francisella novicida (<i>F. novicida</i>)</i>. However, the precise regulatory mechanisms controlling LAP remain poorly understood. Here, we identify enkurin domain-containing protein 1 (ENKD1) as a key negative regulator of LAP during infection with these pathogens. Macrophages infected with <i><i>L.</i> monocytogenes</i> (10403S), <i>S. typhimurium</i> (ATCC14028), or <i>F. novicida</i> (U112) exhibit significant ENKD1 downregulation. ENKD1-deficient macrophages display enhanced antibacterial activity, characterized by increased LAP, higher reactive oxygen species production, enhanced LC3 lipidation on phagosomes, and improved phagosome-lysosome fusion. In vivo, ENKD1<i>-</i>deficient mice <i>ex</i>hibited improved bacterial clearance in the liver and spleen, with notable survival benefits. Mechanistically, ENKD1 interacts with the E3 ubiquitin ligase tripartite motif-containing protein 21 (TRIM21), which mediates degradation of Run domain Beclin-1-interacting and cysteine-rich domain-containing protein (RUBCN) through K48-linked polyubiquitination, thereby dampening RUBCN's role in LAP. Our findings reveal an ENKD1-TRIM21-RUBCN axis that suppresses LAP, providing insights into antibacterial immune regulation and suggesting potential therapeutic strategies to enhance host defense against intracellular pathogens.