A TRAF-like E3 ubiquitin ligase TrafE coordinates ESCRT and autophagy in endolysosomal damage response and cell-autonomous immunity to Mycobacterium marinum

Abstract

Cells are perpetually challenged by pathogens, protein aggregates or chemicals, that induce plasma membrane or endolysosomal compartments damage. This severe stress is recognised and controlled by the endosomal sorting complex required for transport (ESCRT) and the autophagy machineries, which are recruited to damaged membranes to either repair or to remove membrane remnants. Yet, insight is limited about how damage is sensed and which effectors lead to extensive tagging of the damaged organelles with signals, such as K63-polyubiquitin, required for the recruitment of membrane repair or removal machineries. To explore the key factors responsible for detection and marking of damaged compartments, we use the professional phagocyte Dictyostelium discoideum. We found an evolutionary conserved E3-ligase, TrafE, that is robustly recruited to intracellular compartments disrupted after infection with Mycobacterium marinum or after sterile damage caused by chemical compounds. TrafE acts at the intersection of ESCRT and autophagy pathways and plays a key role in functional recruitment of the ESCRT subunits ALIX, Vps32 and Vps4 to damage sites. Importantly, we show that the absence of TrafE severely compromises the xenophagy restriction of mycobacteria as well as ESCRT-mediated and autophagy-mediated endolysosomal membrane damage repair, resulting in early cell death.

The evolutionarily conserved PRP4K-CHMP4B/vps32 splicing circuit regulates autophagy

“…3, 5 and 8). These data are consistent with and extend the finding that Dictyostelium Vps32 is required for autophagosome maturation 57 , and provide a mechanism by which disruption of autophagy impairs development in Dictyostelium by reducing secretion of c-di-GMP. Most astonishingly, we found that the regulatory Prp4k- vps32 splicing circuit is evolutionarily conserved in humans, where mis-splicing of CHMP4B after PRP4K depletion (Fig.…”

“…Previous work combined with the current study, implicates PRP4K in a cluster of cellular functions that are seemingly unrelated, including endocytic trafficking 22 , mitosis/cytokinesis 23,34 , and autophagy in human cells and Dictyostelium. However, these represent a very similar cluster of pathways known to be regulated by ESCRT machinery, including endocytic trafficking, cell abscission and autophagy [54][55][56][57][58] . Given this similarity, we hypothesized that expression of ESCRT genes may be affected by the loss of PRP4K.…”

Flagellin O-linked glycans are required for the interactions between Campylobacter jejuni and Acanthamoebae castellanii

“…Dictyostelium discoideum Ax2(Ka) wild-type cells were grown in axenic conditions at 22 °C in HL5c medium (Formedium) including glucose supplemented with vitamins and micro-elements, as described previously [ 12 ].…”

Temporal genome-wide fitness analysis of Mycobacterium marinum during infection reveals the genetic requirement for virulence and survival in amoebae and microglial cells

“…1A ) ( 3 , 22 , 23 ). D. discoideum has emerged as a powerful genetically and biochemically tractable model to study processes of M. marinum infection, and in particular has recently been used to demonstrate the role of the ESCRT and autophagy machinery in MCV membrane repair ( 24 , 25 ), the role of metal poisoning in bacterial growth restriction ( 26 , 27 ) as well as to identify anti-infective compounds ( 28 , 29 ). Therefore, we use D. discoideum and M. marinum as a model system to study cell-autonomous defense mechanisms that are relevant to the pathogenesis of tuberculosis ( 3 , 8 , 9 ).…”