Development of mouse models for autoinflammatory rare diseases

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 DESCRIPTION (provided by applicant): Immunoproteasomes (i-proteasomes) are an inducible type of proteasomes that have proteolytic activities different from standard proteasomes. They play important roles not only in antigen presentation but also in removal of oxidized proteins that accumulate in stress. They also regulate cell signaling, inflammation and cytokine secretion in a variety of cell types. Recent homozygosity mapping and exome sequencing analysis revealed that one of the i-proteasomal beta subunits, LMP7 (Psmb8 gene product), is mutated in multiple autoinflammatory rare diseases, such as Nakajo-Nishimura syndrome (amyotrophy-fat tissue anomaly), CANDLE syndrome, and an autosomal-recessive autoinflammatory syndrome characterized by joint contractures, muscle atrophy, microcytic anemia, and panniculitis-induced lipodystrophy. There are multiple point mutations of LMP7 identified in European, United States and Asian populations that show similar disease phenotypes. A common feature of the diseases is deformation and dys-function of fat tissue along with problems in other tissues, such as bone and muscle. Although such a point mutation could cause problems, it is critical to recapitulate the disease conditions using mouse models to address whether or how the mutations cause the disease states. Our preliminary study showed that LMP7 deficiency suppresses fat accumulation in the body and fat mobilization in response to fasting and impairs glucose homeostasis. We also found that LMP7 knockdown suppresses adipogenesis in 3T3-L1 cells. Although the preliminary results greatly contributed to our understanding of the function of LMP7 in the disease-related phenotypes, the KO mice are not seen ideal to recapitulate the human disease conditions. The objective of this grant is to establish mouse models that can closely recapitulate the autoinflammatory rare syndromes caused by LMP7 mutation. The genetic mouse models reproducing the disease mutation will be important to decipher disease mechanisms and for development of therapeutic interventions. We will generate mice that harbor a point mutation Thr75Met in LMP7. We will use TALEN-assisted gene targeting technique combined with a donor oligonucleotide to introduce the point mutation into the mouse genome. We demonstrated that T75M mutation is critical for adipocyte differentiation and the i-proteasomal activity in 3T3-L1 adipocytes, indicating that T75M mutation has functional and phenotypic impact in murine cells. To further decipher the pathophysiology of the diseases, we will also use a tissue-specific mouse model and determine the roles of i-proteasomes in macrophages and adipocytes, the two major cell types residing in adipose tissue, in the disease phenotypes. For this, we have already made LMP7 flox mice. The mouse models are unique resources that will greatly contribute to understanding the pathophysiology of the rare diseases. They will also be useful to explore many human diseases belonging to the categories of autoimmune diseases as well as metabolic diseases, cancer and neurodegenerative diseases. They will be essential for therapeutic drug development for the rare diseases.