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Mechanisms of linear ubiquitination by the LUBAC ligase complex

Linear ubiquitin chains are essential regulators of multiple immune signalling cascades. The LUBAC complex is the only known ubiquitin ligase complex capable of assembling these chains. However, it is unclear how LUBAC is regulated, activated, or recruited to signalling complexes. Furthermore, the mechanisms through which linear ubiquitin chains influence the activity of the substrates they are conjugated to are unknown.

Our group is using a multidisciplinary approach combining biochemistry, cellular and chemical biology to unravel the mechanisms underlying linear ubiquitination. We are specifically interested in understanding how linear ubiquitin chains mediate the activation of enzymatic substrates, and the mechanisms in regulating the activity of LUBAC. Through these studies, we aim to unravel the mechanistic role of linear ubiquitin chains in immune signalling and develop experimental strategies that can be universally applied to study ubiquitination.

Linear ubiquitination in immune responses

In mammals, linear ubiquitination plays an important role in the immune responses and cell death. In human patients who suffer from autoimmune disease and myopathy were found to have mutations in genes responsible for the LUBAC components. In mouse models in which LUBAC component is deficient, apoptosis is abnormally increased. For example, apoptosis in multiple organs is increased in deficient mice of a LUBAC component Sharpin. By contrast, knockout of the LUBAC component, HOIP or HOIL-1L in mice leads to embryonic lethality accompanied by increased apoptosis in various tissues. While the involvement of LUBAC-dependent linear ubiquitination in the regulation of apoptosis and immune responses is clear, it remains to be understood how each LUBAC component regulates these events. Moreover, the roles of LUBAC in biology besides inflammatory responses and cell death remain largely elusive. We aim to understand the biological roles of linear ubiquitination using biochemical and cellular techniques coupled with genetically modified mouse models.

Biology of linear ubiquitination using fly models

Recently, we identified the first linear ubiquitination enzyme in Drosophila melanogaster and named it LUBEL (linear ubiquitin E3 ligase). By introduction of mutations at the catalytic domain of LUBEL in flies, we established linear ubiquitin chain-deficient fly models, which enable us to explore the different aspects of linear ubiquitination in a whole organism. These LUBEL mutant flies are viable and found to be defective in heat tolerance accompanied with mobility disability. Our goal is to understand the fundamental biological functions of linear ubiquitination using a newly established fly models, especially the roles outside of inflammatory response. 

Ubiquitination and ubiquitin enzymes in autophagy

Autophagy is a pivotal lysosomal degradation pathway that protects cells from nutrient deprivation, as well as several other stress conditions. To ensure rapid activation, but at the same time to avoid degradation of essential material, it needs to be tightly controlled. One key mechanism to regulate autophagy is posttranslational modification of autophagy players with the small protein ubiquitin, ubiquitination. Even though ubiquitination is crucial for autophagy regulation, to date, only a handful of ubiquitin enzymes including LUBAC have been identified and characterized to regulate autophagy. In this project, we aim to characterize ubiquitin enzymes and their role in autophagy. In this way, we aim to broaden the current knowledge of how ubiquitination contributes to autophagy regulation.