High-affinity binding of the trimeric fibre protein to a cell surface primary receptor is a common feature shared by all adenovirus serotypes. Recently, a long elusive species B adenovirus receptor has been identified. Desmoglein 2 (DSG2) a component of desmosomal junction, has been reported to interact at high affinity with Human adenoviruses HAd3, HAd7, HAd11 and HAd14. Little is known with respect to the molecular interactions of adenovirus fibre with the DSG2 ectodomain. By using different DSG2 ectodomain constructs and biochemical and biophysical experiments, we report that the third extracellular cadherin domain (EC3) of DSG2 is critical for HAd3 fibre binding. Unexpectedly, stoichiometry studies using multi-angle laser light scattering (MALLS) and analytical ultra-centrifugation (AUC) revealed a non-classical 1:1 interaction (one DSG2 per trimeric fibre), thus differentiating ‘DSG2-interacting’ adenoviruses from other protein receptor interacting adenoviruses in their infection strategy.

Cells are perpetually challenged by pathogens, protein aggregates or chemicals, that induce plasma membrane or endolysosomal compartments damage. Endolysosomal perforations are recognised as severe stress, however the mechanisms of the cellular response that ensure quality control, repair and endolysosomal homeostasis are just beginning to be unravelled. The endosomal sorting complex required for transport (ESCRT) and the autophagy machinery are recruited to damaged membranes to either repair or to remove membrane remnants. Crucial element of the endolysosomal damage response (ELDR) are factors that sense damage, paralleled by extensive tagging of the damaged organelles with signals, such as ubiquitin, required for the recruitment of ELDR components. Unattended membrane damage leads to leakage of harmful components including protons and reactive oxygen species that cause cell death. To explore ELDR 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 components. Importantly, we show that the absence of TrafE severely compromises the xenophagy restriction of bacteria as well as autophagy-mediated and ESCRT-mediated ELDR, resulting in early cell death.

No 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 recombinant antibodies AI842, AI843, AI844 and AI177 failed to detect by immunofluorescence a C-terminally FLAG-tagged LmpA fusion protein expressed in Dictyostelium discoideum.