Kile Lab research
Our lab has a longstanding interest in haematopoiesis and cell death. We focus particularly on the megakaryocyte lineage, from stem cells through to platelets. Using primarily molecular approaches, we seek to understand the role of pathways such as apoptosis, pyroptosis and necroptosis in haematopoietic biology. This includes elucidating pathways at steady state, and in disease settings such as leukaemia and inflammatory disease. Our work extends to the development of new chemical probes to study cell death in the haematopoietic system and other tissues.
We are researching the molecules that underpin the formation and function of blood cells. Our major interests are:
- The regulation of fundamental processes, such as cell death.
- How these processes contribute to the development of leukaemia and inflammatory disease.
- Identifying new strategies to target these processes as treatments for diseases.
1. Apoptosis, mitochondrial damage and the innate immune response
We recently demonstrated that, during apoptosis, mitochondria undergo herniation. Large macropores form in the outer membrane, allowing the inner membrane to balloon out into the cytoplasm, bringing mtDNA with it. This leads to activation of the cGAS/STING innate immune signalling pathway and the production of inflammatory cytokines like interferon beta. Published in Science (McArthur et al. 2018) this is the first documented example of mtDNA escape. Our work raises questions about the relationship between apoptosis, mtDNA and inflammatory responses. mtDNA is found in the serum of healthy people, and elevated levels have been reported in a range of autoinflammatory and autoimmune disorders. This is thought to be the result of aberrant apoptosis or failed apoptotic cell clearance. We are now examining the requirement for apoptotic processes in the generation of extracellular mtDNA.
Lattice light-sheet imaging of cells undergoing apoptosis, the mitochondria (red: TOMM20-Halo) lose their elongation and become circular, then mtDNA (green: TFAM-mNeonGreen) escapes. We have termed this process “mitochondrial herniation”. Images were captured at a rate of one 3D volume every 8 seconds.
Single mitochondria undergoing nucleoid externalization (red: TOMM20-Halo, green, TFAM-mNeonGreen, blue: mRuby2-BAX). 3D surface reconstruction overlaid upon original LLSM data with Imaris software.Fig 4E, McArthur et al. Science 2018 Vol 359
Reconstructed cryo-tomogram of an apoptotic herniating mitochondria moving through the z axis. McArthur et al, Science 2018 Vol 359
2. The role of senescence, death and clearance in blood cell homeostasis
The ability of the adaptive immune system to mount an antibody response against virtually any pathogen relies on a diverse B cell repertoire, and on the continuous generation of newly generated B cells in the bone marrow. It is well established that the number of B cells produced in the bone marrow diminishes with age. This is thought to limit the effectiveness with which the elderly mount protective antibody responses. The mechanisms underpinning this phenomenon are still obscure. We have generated a novel model of immune senescence. Using gene expression profiling, cell biology and genome editing techniques, we are exploring the relationship between cell death, normal homeostasis and senescence, and how perturbations in these processes might contribute to disease.
Apoptotic Tango: megakaryopoiesis in mice with a megakaryocyte-specific deletion of Mcl-1 (Mcl-1Pf4Δ/Pf4Δ) is normal. However, 2.5 hours after treatment with the BH3 mimetic ABT-737, megakaryocytes from these mice exhibit hallmarks of apoptosis with nuclear pyknosis and cytoplasmic dysmorphology evident. The image shows two megakaryocytes undergoing “a dance of death” in the bone marrow of Mcl-1Pf4Δ/Pf4Δ mice, indicating essential roles for both Bcl-xL and Mcl-1 in the viability of the megakaryocyte lineage. Debrincat et al. Blood 2012 Vol 119
B cell development: (A) Schematic summarising the phenotypic markers used to identify B cells at different stages of development, shown together with the recombination events occurring at each stage. (B) Flow cytometric analysis of developing B cells.
3. Caspases, infection and cancer
Caspases are enzymes that are activated during programmed cell death (apoptosis) to quickly dismantle the dying cell. We have shown that, in addition to hastening cellular demise, caspases prevent dying cells from triggering the innate immune system (White, Cell 2014). This raises interesting questions about how caspases facilitate embryonic development, and how they function in disease settings such as viral infection and cancer. We are utilising genetic and cell biology approaches to unravel these processes, and exploring whether drugs that inhibit caspases might have clinical potential.
Activation of Bak and Bax, essential mediators of apoptosis, triggers mtDNA release, which in turn stimulates the cGAS/STING pathway to induce IFN-β production. Activation of the apoptotic caspase cascade blocks the cGAS/STING pathway. Inhibition or genetic deletion of caspases causes apoptotic cells to secrete IFN-β. White et al. Cell 2014 Vol 159
We collaborate with many scientists and research organisations around the world. Click on the map to see the details for each of these collaborators (dive into specific publications and outputs by clicking on the dots).
Student research projects
The Kile Lab offers a variety of Honours, Masters and PhD projects for students interested in joining our group. There are also a number of short term research opportunities available.
Please visit Supervisor Connect to explore the projects currently available in the Kile Lab.