Actin Cytoskeleton Remodelling in Apoptosis-induced Proliferation and Tissue Growth Control

In multicellular organisms including humans, damaged or stressed cells are removed by apoptosis, a programmed cell death pathway, to prevent their detrimental effects on the hosts. Studies in the past decade have revealed that dying cells, before they are removed, can signal their neighbouring surviving cells to induce extra cell divisions and production of new cells which compensates for the loss of dead cells. This process, critical for balancing cell numbers and maintenance of tissue homeostasis, is termed as Apoptosis-induced Proliferation (AiP). Uncontrolled AiP contributes to the development of diseases including cancer. It is therefore important to understand how AiP is controlled at the cellular and molecular level. By using Drosophila as a genetic model organism, we observed an increase of actin filaments, fundamental components of the cellular scaffolding structure - cytoskeleton, in cells emitting AiP signals. Interestingly, such an increase of actin filaments, i.e. actin remodelling, is required for production of reactive oxygen species (ROS) and activation of c-Jun N-terminal kinase (JNK) signalling, two key processes known to trigger AiP. The proposed work is to determine how actin remodelling is regulated in apoptotic dying cells and how it controls ROS production and JNK activation leading to AiP. We will also examine roles of actin remodelling in the development of malignant tumour. Dissecting the molecular control of actin remodelling and its cellular functions in both AiP and tumourigenesis will significantly impact our understanding of physiological tissue recovery and pathological tumour development.

Apoptosis-induced Proliferation (AiP) is a compensatory mechanism critical for the maintenance of tissue homeostasis in response to stress-induced apoptotic cell loss. Studies in multiple organisms have revealed that AiP is an evolutionarily conserved phenomenon relevant to tissue regeneration and, under pathological conditions, it contributes to tumourigenesis. Therefore, understanding of the molecular mechanisms controlling AiP has therapeutic implications. Intriguingly, apoptotic cells activate AiP through a caspase-driven signalling cascade. In Drosophila epithelial cells, the Caspase 9 ortholog Dronc initiates AiP via an increased production of reactive oxygen species (ROS) and activation of c-Jun N-terminal kinase (JNK) signalling. However, the specific mechanisms connecting these key factors of AiP remain unknown. We recently identified F-actin polymerisation as a critical mediator of AiP acting downstream of Dronc but upstream of ROS and JNK. In the proposed project, we will elucidate: 1) how Dronc promotes F-actin polymerisation; 2) how F-actin polymerisation regulates production of ROS and activation of JNK; and 3) how F-actin polymerisation controls the neoplastic tumour growth. Answers to these questions will provide novel insights into the cellular signalling processes that trigger AiP and their important roles in tumourigenesis.

BBSRC STRATEGIC PRIORITIES
Due to the broad relevance of Apoptosis-induced Proliferation (AiP) to tissue regeneration and human pathological conditions including cancer, this basic science research proposal falls within the BBSRC's strategic area of "world-class bioscience", specifically, "driving advances in fundamental bioscience for better health". It also meets BBSRC's strategic priorities on "Healthy ageing across the lifecourse" as it is "generating new knowledge to advance regenerative biology" and "The replacement, refinement and reduction (3Rs) in research using animals" because we use Drosophila to replace the use of mammals and address a research topic directly relevant to humans. Our research outcomes may contribute to development of regenerative medicine and novel cancer therapies. Moreover, the proposed project will have direct impacts on developing innovative research tools, advancing cutting-edge research and training skilled researchers which all contribute to BBSRC's strategic plan on "maintaining the UK's position as a global leader".

INDUSTY AND CLINICAL APPLICATIONS
Understanding the molecular mechanisms underlying AiP can provide potential drug targets for regenerative medicine and cancer therapy. Specifically, outcomes of this research are to elucidate how actin cytoskeleton remodelling is activated in AiP and how production of reactive oxygen species (ROS) and c-Jun N-terminal kinase (JNK) are activated by this process. Notably, actin cytoskeleton remodelling, ROS and JNK have all been recognised as contributing factors to tissue regeneration and cancer development. Our research will establish a connection among these factors in vivo in an intact model organism. Therefore, in addition to providing disease-relevant information, the Drosophila assays employed in the project can also be potentially applied to clinical and industrial uses e.g. as low-cost and alternative tools for drug screen or validation of signalling responses in vivo.

ACADEMIA AND SCIENTIFIC COMMUNITY
The proposed research will employ Drosophila as a model organism to study roles of actin cytoskeleton remodelling in activation of AiP. It will investigate caspase functions, actin cytoskeleton regulators, stress response signalling molecules and their implications in tumourigenesis. Therefore, the proposed work will be of interest to a wide range of research fields including protease function, cytoskeleton dynamics, stress response signalling transduction, cell death and cancer biology. Moreover, the assays developed and used in the project will contribute to the scientific community as valuable research tools. These tools provide not only scientific benefits but also the possibilities to reduce and replace the use of mammals in cancer research and drug development.

STUDENTS AND PUBLIC
The proposed research employs genetically amenable model organisms to study cellular processes and signalling pathways relevant to human diseases. This concept as well as knowledge and skills required to conduct Drosophila genetics, microscopy imaging, statistical analysis, molecular and biochemical assays will be passed to all the researchers, who will be involved in the project, through regular meetings, hands-on teaching and established collaborations. We will also communicate with the general public and engage school students by presenting our research at the University Community Days and other research showcase events. This is to enhance public awareness and understanding of our research as well as its potential therapeutic benefits.