Proteins play central roles in cellular processes, and cells are equipped with quality control mechanisms to ensure they function properly. There are mainly two kinds of protein quality control systems in a cell: a molecular chaperone system which helps proteins fold correctly, and a degradation system which facilitates protein disposal. When quality control mechanisms fail, cells become susceptible to various stresses, and diseases such as neurodegenerative disorders result. In our laboratory, we are investigating mechanisms of protein quality control during cellular stress. We use budding yeastSaccharomyces cerevisiae since it provides an advantageous experimental system for elucidating molecular mechanisms due to the ease of genetic manipulation.
1 Analysis of heat shock response
Organisms try to survive in various stressful environments. For example, when exposed to high temperatures they increase the expression of various molecular chaperone and ubiquitin proteins to protect or maintain proteins from unfolding, or target unfolded proteins for degradation. In our laboratory, we are investigating molecular mechanisms of chronic heat stress. We analyze proteins which are specifically induced by the chronic stress, and investigate the role of ubiquitination in these processes. In addition, we analyze changes of cellular structures by the stress. In the future, we hope to understand how organisms respond against global warming at the molecular level.
2 Ubiquitin homeostasis and stress response
Within a cell, the ‘ubiquitin homeostasis’ process functions to tightly regulate ubiquitin levels through several different ways. This is necessary because ubiquitin is an orchestrator of a remarkable array of cellular tasks. For example, when this tiny protein “tag” is attached to a target the resultant complex is earmarked for degradation in the proteasome. Alternatively, the complex may be transported to a specific cellular compartment such as the plasma membrane or endosome. A large amount of ubiquitin is required to enable normal protein quality control. Ubiquitin is encoded by several genes, and is also directly recycled by the cell, but interestingly, it is not in excess. We have discovered that one additional way in which ubiquitin levels are regulated is through the balanced activity of a de-ubiquitinating enzyme and an inhibitor. We are working to understand and elucidate additional ubiquitin homeostasis mechanisms.
3 Analysis of ubiquitin-related chaperone VCP/Cdc48
VCP/Cdc48 is a member of the AAA + (ATPase associated with diverse cellular activities) family of proteins. Within the cell, VCP/Cdc48 is abundant and ubiquitously distributed. VCP/Cdc48 participates in diverse cellular activities, including ubiquitin-proteasome-mediated protein degradation, fusions to the endoplasmic reticulum (ER) and Golgi membranes, autophagy, and endolysosomal sorting. There are many different single amino acid-substitutions in VCP that have been reported to cause Inclusion Body Myopathy associated with Paget Disease of bone and Frontotemporal Dementia (IBMPFD), an autosomal dominant inherited human disease, and familial amyotrophic lateral sclerosis (ALS). To understand how these mutant proteins cause disease such as IBMPFD and ALS, we express human pathogenic VCP variants in yeast and observe resultant cellular effects. In addition, we are conducting genetic suppressor and enhancer screens to look for modifiers of VCP/Cdc48 function.
Yeast Saccharomyces cerevisiae