Sqstm1-GFP knock-in mice reveal dynamic actions of Sqstm1 during autophagy and under stress conditions in living cells.
Sqstm1 serves as a signaling hub and receptor for selective autophagy. Consequently, dysregulation of Sqstm1 causes imbalances in signaling pathways and disrupts proteostasis, thereby contributing to the development of human diseases. Environmental stresses influence the level of Sqstm1 by altering its expression and/or autophagic degradation, and also changes the localization of Sqstm1, making it difficult to elucidate the actions and roles of this protein. In this study, we developed knock-in mice expressing Sqstm1 fused to GFP (Sqstm1-GFP(KI/+)). Using these Sqstm1-GFP(KI/+) mice, we revealed for the first time the dynamics of endogenous Sqstm1 in living cells. Sqstm1-GFP was translocated to a restricted area of LC3-positive structures, which primarily correspond to the inside of autophagosomes, and then degraded. Moreover, exposure to arsenite induced expression of Sqstm1-GFP, followed by accumulation of the fusion protein in large aggregates that were degraded by autophagy. Furthermore, suppression of autophagy in Sqstm1-GFP(KI/+) mouse livers caused accumulation of Sqstm1-GFP and formation of GFP-positive aggregate structures, leading to severe hepatic failure. These results indicate that Sqstm1-GFP(KI/+) mice are a useful tool for analyzing Sqstm1 in living cells and intact animals.
Department of Biochemistry, Niigata University Graduate School of Medical and Dental Sciences, Chuo-ku, Niigata 951-8510, Japan Division of Clinical Nephrology and Rheumatology, Niigata University Graduate School of Medical and Dental Sciences, Chuo-ku, Niigata 951-8510, Japan.
Atsushi
A
Eino
Department of Biochemistry, Niigata University Graduate School of Medical and Dental Sciences, Chuo-ku, Niigata 951-8510, Japan.
Shun
S
Kageyama
Department of Anatomy and Histology, Fukushima Medical University School of Medicine, Hikarigaoka, Fukushima 960-1295, Japan.
Takefumi
T
Uemura
Department of Anatomy and Histology, Fukushima Medical University School of Medicine, Hikarigaoka, Fukushima 960-1295, Japan.
Hiromichi
H
Annoh
Department of Biochemistry, Niigata University Graduate School of Medical and Dental Sciences, Chuo-ku, Niigata 951-8510, Japan.
Tetsuya
T
Saito
Division of Clinical Nephrology and Rheumatology, Niigata University Graduate School of Medical and Dental Sciences, Chuo-ku, Niigata 951-8510, Japan.
Ichiei
I
Narita
Department of Anatomy and Histology, Fukushima Medical University School of Medicine, Hikarigaoka, Fukushima 960-1295, Japan komatsu-ms@med.niigata-u.ac.jp waguri@fmu.ac.jp.
Satoshi
S
Waguri
Department of Biochemistry, Niigata University Graduate School of Medical and Dental Sciences, Chuo-ku, Niigata 951-8510, Japan komatsu-ms@med.niigata-u.ac.jp waguri@fmu.ac.jp.
Masaaki
M
Komatsu
D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D001343:Autophagy; D005786:Gene Expression Regulation; D055879:Gene Knock-In Techniques; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D006360:Heat-Shock Proteins / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D017093:Liver Failure / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D010588:Phagosomes / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D000071456:Sequestosome-1 Protein; D013312:Stress, Physiological
D048868:Adaptor Proteins, Signal Transducing; D006360:Heat-Shock Proteins; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D011993:Recombinant Fusion Proteins; D000071456:Sequestosome-1 Protein; C101343:Sqstm1 protein, mouse; D049452:Green Fluorescent Proteins
Autophagy; Knock-in mice; Nrf2; Selective autophagy; Sqstm1