Oral Presentation 49th Annual Scientific Meeting of the Australian and New Zealand Society for Immunology 2021

Using macrophage heterogeneity and single-cell RNA sequencing to characterize molecular mechanisms of the zinc toxicity antimicrobial response (#80)

Jessica B von Pein 1 2 , Claudia J Stocks 1 2 , Stacey B Andersen 3 4 , Jon Xu 3 4 , Nicholas D Condon 1 , Minh Duy Phan 5 6 , Kate Peters 5 6 , Ronan Kapetanovic 1 2 5 , Mark Schembri 5 6 , Matt Sweet 1 2 5
  1. Institute for Molecular Bioscience (IMB), University of Queensland, St Lucia, Queensland, Australia
  2. IMB Centre for Inflammation and Disease Research, University of Queensland, St Lucia, Queensland, Australia
  3. Genome Innovation Hub, University of Queensland, St Lucia, Queensland, Australia
  4. UQ Sequencing Facility, Brisbane
  5. Australian Infectious Diseases Research Centre, University of Queensland, St Lucia, Queensland, Australia
  6. School of Chemistry and Molecular Biosciences, University of Queensland, St Lucia, Queensland, Australia

Urinary tract infections are one of the most common bacterial infections in humans, with uropathogenic E. coli (UPEC) being the primary cause of these infections. Understanding effective host defence pathways could facilitate the development of new therapeutics to treat UPEC infections. During infection, innate immune cells, including macrophages, can manipulate nutrient availability to either starve or toxify microorganisms with trace elements as opposing antimicrobial strategies. Macrophages deploy zinc toxicity against intracellular bacteria, but molecular mechanisms underlying this pathway are poorly understood. Here, we investigate the mechanisms of the macrophage zinc toxicity response in human monocyte-derived macrophages (HMDM). Using Fluozin-3 staining and microscopy, we found that CSF-1-derived and GM-CSF-derived HMDMs exhibit different basal zinc distributions. Through assessment of mammalian and bacterial gene expression, as well as intramacrophage bacterial survival, we demonstrate that both macrophage populations engage zinc toxicity and zinc starvation; however, CSF-1-derived HMDMs are skewed towards toxicity, whereas GM-CSF HMDMs are skewed towards zinc starvation responses. Using an E. coli strain that reports zinc stress, we demonstrate variation of zinc toxicity responses between individual cells within CSF-1-derived HMDM. To further delineate the heterogeneity of macrophage zinc toxicity responses, we developed a dual single-cell RNA sequencing platform to assess both host and pathogen transcriptomes associated with the zinc toxicity response in HMDM. This unique approach allows us to match individual HMDM transcriptomes with those of zinc-stressed bacteria, thus revealing host genes that may contribute to the zinc toxicity response. Ongoing studies aim to functionally assess gene candidates from this approach as mediators of the macrophage zinc toxicity response. Such knowledge may ultimately guide the development of host-derived therapies for the treatment of antibiotic-resistant bacterial infections.