![An inverse relationship between PLD2 and UCP1 expression in adipose tissues upon HFD feeding. (A) Quantitative RT-PCR of Pld1 and Pld2 mRNA expression in various adipose tissues from 8-wk-old WT mice (n = 6/group). (B) Representative Western blot images of PLD1, PLD2, UCP1, and GAPDH in BAT from HFD (60% fat)-fed WT mice for 0, 4, and 12 wk at 22°C. (C) Representative Western blot images of PLD2, UCP1, and GAPDH in BAT from NCD or HFD (45% fat)-fed WT mice for 4 wk at 30°C. (D) Experiments to test the effects of sympathetic nerve system stimulation on the level of PLD2. Schematic illustration depicting WT mice incubated at 4°C for 1 wk or injected with the β3AR agonist, CL, at 1 mg/kg for 7 consecutive days (left). Representative Western blot images of PLD2, UCP1, GAPDH, and β-Actin (middle), and quantification of PLD2 protein level (right) in BAT and ingWAT (n = 6/group). (E) Representative Western blot images of PLD2, UCP1, and GAPDH in BAT and ingWAT from vehicle- or CL-stimulated mice at 22°C or 30°C for 1 wk. (F) PLD activity levels in lysates of BAT and ingWAT from 8-wk-old WT mice incubated at 4°C for 1 wk or injected with CL at 1 mg/kg for 7 consecutive days (n = 6/group). (G) The proteasomal activity in the lysates of ingWAT and BAT from NCD or HFD (45% fat)-fed WT mice for 4 wk at 30°C (n = 5/group). (H) Representative Western blot images of PLD2, UCP1, and β-Actin (left) and quantification of PLD2 (right) in isoprenaline (1 µM)- and SR59230A (1 µM)-treated primary brown adipocytes. (I) Representative Western blot images of PLD2 and β-Actin in isoprenaline (1 µM)-, bortezomib (20 nM)-, and chloroquine (10 µM)-treated primary brown adipocytes. (J) The proteasomal activity in the lysates of ingWAT and BAT from HFD (60% fat)–fed WT mice for 0, 4, and 12 wk at 22°C (n = 5/group). The data are presented as mean ± SEM. *, P < 0.05; **, P < 0.01; ***, P < 0.001 by two-tailed Student’s t test (A, D [right], F, G, H [right], and J). Data are representative of at least two (C, E, H, and I) or three independent experiments (A, B, D, F, G, and J). a.u., arbitrary unit; Veh, vehicle.](https://cdn.rupress.org/rup/content_public/journal/jem/219/2/10.1084_jem.20211523/3/jem_20211523_fig1.png?Expires=1767722027&Signature=xi7Bdijq81EGl8xLpcKZIbeTSLFqYZriAew2zTFiv9LC~lzH~wNVO3EG6OeugiNh-p7i0IG~pMc8IDO2vvYD03OyFgQVQBhKtT8CGGb9TZ11yaYMQuvZdE4VEGFJDfjNk92NpM9Cy798RXEDHw77IJKz8-WsUq1JMleKJFTUbIBfEdbS~u~nY0MSJVntEDz75~RfAErNy0QuHfgclgYsWwy91JNKXU3mKJu1ezoBZQ4BdYSnzdNHPIX4kMm6UxWaqpjvbqKjtNbLu7uNwh64dfpwfjih7wzNlKIY4bUYg4kfYY-gSOKTxZkHKFCk~GpEEP-Q2MxbUMyX7YiO3N9Ryw__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
An inverse relationship between PLD2 and UCP1 expression in adipose tissues upon HFD feeding. (A) Quantitative RT-PCR of Pld1 and Pld2 mRNA expression in various adipose tissues from 8-wk-old WT mice (n = 6/group). (B) Representative Western blot images of PLD1, PLD2, UCP1, and GAPDH in BAT from HFD (60% fat)-fed WT mice for 0, 4, and 12 wk at 22°C. (C) Representative Western blot images of PLD2, UCP1, and GAPDH in BAT from NCD or HFD (45% fat)-fed WT mice for 4 wk at 30°C. (D) Experiments to test the effects of sympathetic nerve system stimulation on the level of PLD2. Schematic illustration depicting WT mice incubated at 4°C for 1 wk or injected with the β3AR agonist, CL, at 1 mg/kg for 7 consecutive days (left). Representative Western blot images of PLD2, UCP1, GAPDH, and β-Actin (middle), and quantification of PLD2 protein level (right) in BAT and ingWAT (n = 6/group). (E) Representative Western blot images of PLD2, UCP1, and GAPDH in BAT and ingWAT from vehicle- or CL-stimulated mice at 22°C or 30°C for 1 wk. (F) PLD activity levels in lysates of BAT and ingWAT from 8-wk-old WT mice incubated at 4°C for 1 wk or injected with CL at 1 mg/kg for 7 consecutive days (n = 6/group). (G) The proteasomal activity in the lysates of ingWAT and BAT from NCD or HFD (45% fat)-fed WT mice for 4 wk at 30°C (n = 5/group). (H) Representative Western blot images of PLD2, UCP1, and β-Actin (left) and quantification of PLD2 (right) in isoprenaline (1 µM)- and SR59230A (1 µM)-treated primary brown adipocytes. (I) Representative Western blot images of PLD2 and β-Actin in isoprenaline (1 µM)-, bortezomib (20 nM)-, and chloroquine (10 µM)-treated primary brown adipocytes. (J) The proteasomal activity in the lysates of ingWAT and BAT from HFD (60% fat)–fed WT mice for 0, 4, and 12 wk at 22°C (n = 5/group). The data are presented as mean ± SEM. *, P < 0.05; **, P < 0.01; ***, P < 0.001 by two-tailed Student’s t test (A, D [right], F, G, H [right], and J). Data are representative of at least two (C, E, H, and I) or three independent experiments (A, B, D, F, G, and J). a.u., arbitrary unit; Veh, vehicle.