Alternative roles for autophagy receptors in host-pathogen interactions – AutoHostPath

Understanding the molecular basis of host-pathogen interactions is critical for the discovery of new therapeutic approaches to treat infectious diseases, which in the face of increasing antibiotic resistance and emerging pathogens, is a major priority medical research in Europe. Studying the signalling pathways in immune cells during infection also gives us fundamental information about the mechanisms of immune cell activation during inflammation that is relevant for other human diseases including autoimmune disease, cardiovascular disease and even cancer.

Macroautophagy is an evolutionarily conserved mechanism for recycling cellular nutrients during starvation through lysosomal degradation of cytoplasmic material, but it also serves as a cell autonomous defence mechanism against intracellular microorganisms. Recent studies suggest that specific components of the macroautophagy machinery may have a wider impact on cell biology and the immune response, including the regulation of signal transduction in innate immune cells. Autophagy receptors recognise substrates for autophagy based on distinctive ‘eat me’ signals, these include the ubiquitin (Ub)-binding proteins; p62, NBR1, NDP52 and OPTN. These proteins share some common features, including Ub-binding domains (UBA) and LC3-interacting domains (LIR), but they also have distinctive properties that have been implicated in cellular process relevant to innate and adaptive immunity – independently of autophagy. In fact, these proteins may directly participate in innate immune signalling pathways and thus act as alternative pathogen-recognition receptors (PRR) for invasive microbes. However, their specific roles in host-pathogen interactions and innate immunity remain to be fully explored. Recent studies have shown that LC3 – the classical marker of macroautophagy – can be recruited to phagosomes during conventional phagocytosis, so called LC3-associated phagocytosis (LAP). The physiological significance of LAP is still unclear, but a recent study showed that LAP was specifically required for induction of type I IFN (IFNa/ß) after phagocytosis of immune complexes containing DNA, suggesting LAP may couple phagocytosis to specific innate immune signalling pathways. However, the role of autophagy receptors during LAP has not been addressed.

Pore-forming toxins (PFTs) are the largest family of bacterial protein toxins and they represent major virulence factors for a number of important human pathogens, including Streptococcus pyogenes (group A Streptococcus ; GAS) and Staphylococcus aureus (SA). Beyond their cytolytic activities at high concentrations, the effects of these toxins on host pathogen interactions are poorly understood. Our previous studies have shown important roles for PFTs in modulation of innate immune signalling pathways in macrophages. Here we present new preliminary data that shows the PFT Streptolysin O (SLO), a critical virulence factor for GAS, promotes recruitment of LC3 during phagocytosis in macrophages, which is also associated with recruitment of the autophagy receptors OPTN, NBR1 and p62. Furthermore, using knock-in mice bearing a mutation in the UBA of OPTN (OptnD477N), we show that Ub-binding by OPTN is required for SLO-dependent induction of IFNß in GAS-infected macrophages. These preliminary data suggest that SLO-mediated OPTN recruitment couples phagocytosis of GAS with specific innate immune signalling pathways that may affect host defence (Fig.4).

In this project will further explore the mechanisms and consequences of OPTN/NBR1/p62 recruitment in macrophages during GAS infection – and investigate their potential roles as novel phagosomal signalling adaptors. We will also extend these studies to another important Gram-positive bacteria, SA – which expresses related toxins Panton Valentine leukocidin (PVL) and alpha-toxin (Hla).