title: Genetically encoded fluorescent thermosensors visualize subcellular thermoregulation in living cells.
abstract: In mammals and birds, thermoregulation to conserve body temperature is vital to life. Multiple mechanisms of thermogeneration have been proposed, localized in different subcellular organelles. However, visualizing thermogenesis directly in intact organelles has been challenging. Here we have developed genetically encoded, GFP-based thermosensors (tsGFPs) that enable visualization of thermogenesis in discrete organelles in living cells. In tsGFPs, a tandem formation of coiled-coil structures of the Salmonella thermosensing protein TlpA transmits conformational changes to GFP to convert temperature changes into visible and quantifiable fluorescence changes. Specific targeting of tsGFPs enables visualization of thermogenesis in the mitochondria of brown adipocytes and the endoplasmic reticulum of myotubes. In HeLa cells, tsGFP targeted to mitochondria reveals heterogeneity in thermogenesis that correlates with the electrochemical gradient. Thus, tsGFPs are powerful tools to noninvasively assess thermogenesis in living cells.
authors.affiliation: 1] Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Kyoto, Japan. [2] Department of Technology and Ecology, Hall of Global Environmental Studies, Kyoto University, Kyoto, Japan. [3] Core Research for Evolution Science and Technology, Japan Science and Technology Agency, Tokyo, Japan.
authors.forename: Shigeki
authors.initials: S
authors.lastname: Kiyonaka
authors.forename: Taketoshi
authors.initials: T
authors.lastname: Kajimoto
authors.forename: Reiko
authors.initials: R
authors.lastname: Sakaguchi
authors.forename: Daisuke
authors.initials: D
authors.lastname: Shinmi
authors.forename: Mariko
authors.initials: M
authors.lastname: Omatsu-Kanbe
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Matsuura
authors.forename: Hiromi
authors.initials: H
authors.lastname: Imamura
authors.forename: Takenao
authors.initials: T
authors.lastname: Yoshizaki
authors.forename: Itaru
authors.initials: I
authors.lastname: Hamachi
authors.forename: Takashi
authors.initials: T
authors.lastname: Morii
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Mori
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D052437:Adipocytes, Brown / Q000166:cytology; D001426:Bacterial Proteins / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000737:chemistry*; D006367:HeLa Cells; D006358:Hot Temperature; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods; D008928:Mitochondria / Q000378:metabolism; D018485:Muscle Fibers, Skeletal / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism; D011487:Protein Conformation; D019779:Salmonella enterica / Q000378:metabolism*; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D018076:DNA, Complementary; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; C075138:TlpA protein, Salmonella