![Supplementation of serotonin restored the systemic UPRmtactivation in tmbim-2 mutants. (A) Representative photomicrographs of dve-1 reporter in animals expressing neuronal Q40 and neurotoxin TeTx in ciliated sensory neurons (ythEx488[xbx-1p::TeTx::sl2::mcherry]). Scale bar, 250 μm. (B) Quantification of the number of intestinal nuclei puncta with GFP signal per worm as shown in A. n ≥ 15 worms. (C) Representative photomicrographs of the ADF neurotransmitter exocytosis reporter (ythEX766[srh-142p-SYP-Phluorin+unc-119(+)]) in control and srh-142p::TeTx::SL2::mCherry expressing animals. For the control group, mCherry channel image was not acquired (N/A) due to the absence of mCherry expression. The imaging used Z-stacks. Scale bar, 10 μm. (D) Representative photomicrographs of hsp-6 reporter (zcIs3[hsp-6p::gfp]) expression in animals treated with vehicle control or 50 mM serotonin (5-HT). (E) Quantification of hsp-6p::gfp expression. The genotypes are as in D. n ≥ 15 worms. (F) Representative photomicrographs of hsp-6 reporter expression in rgef-1p::Q40::yfp; tmbim-2(yth26) animals treated with vehicle control or 50 mM 5-HT. (G) Quantification of hsp-6p::gfp expression. The genotypes are as in F. n ≥ 15 worms. (H) Representative TEM images of synaptic profiles, including WT, tmbim-2(yth57), and tmbim-2(yth26). Arrows indicate active zone; Scale bar, 200 nm. (I) Quantification of the number of synaptic vesicles at the active zone as shown in H. n ≥ 5 worms. (J) Quantification of the size of synaptic vesicles at the active zone as shown in H. n ≥ 5 worms. (K) Dot plot shows the average expression of Tmbim2 in 17 mouse organs or tissue at the age of 3 mo. WBC, white blood cells; BAT, brown adipose tissue; SCAT, subcutaneous adipose tissue; GAT, gonadal adipose tissue; MAT, mesenteric adipose tissue. (L) Representative confocal photomicrograph of animals spermatheca expressing TMBIM-2 at different ages. The imaging used z-planes. Scale bar, 20 μm. (M) Quantitative analysis of TMBIM-2::GFP protein levels in ROI shown in L. n ≥ 8 worms. (N) Survival analysis of WT (blue) and tmbim-2 (yth26) (yellow); n ≥ 80 worms. (O) Survival analysis of control (blue) and ADF neuron overexpressing TMBIM-2 (yellow); n ≥ 80 worms. ***P < 0.001, **P < 0.01, ns denotes P > 0.05 via unpaired two-tailed Student’s t test. In N and O, we used The Log-rank (Mantel-Cox) test for statistical analysis. Error bars, SEM.](https://cdn.rupress.org/rup/content_public/journal/jcb/224/5/10.1083_jcb.202408050/2/jcb_202408050_figs5.png?Expires=1770186634&Signature=yDD2Y0l6~KsKY47lGSzO3KmbXGXJTAVPi3MZd9dTFavQ7Zf5AusyoUeqLVuscrTM8-EwsrXBOvwhQV73TgFRfaP3o8rlGPDucM5oYhbVzGifKYma95aoWPkCxFMk9Upp~fvT1GPaemD0Ew7SwhySsVfAt224wUxlnXUSILqEoJZIF~2k7TuksTHIbytIb1YJ46cdF5HZhbwo800VIVUkXPvB0zBa-16RfYGgLuOiMQc56EoHoVPF2vBZcHkTST0UPCe96MqPkpaN4QSjRuuYMzioWM8bxwLRxDSpEQzrtebqAkbBvHX3h-S4hwXO-yWFWpVJ7ygNzNLon4NwrRxZng__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Supplementation of serotonin restored the systemic UPR mt activation in tmbim-2 mutants. (A) Representative photomicrographs of dve-1 reporter in animals expressing neuronal Q40 and neurotoxin TeTx in ciliated sensory neurons (ythEx488[xbx-1p::TeTx::sl2::mcherry]). Scale bar, 250 μm. (B) Quantification of the number of intestinal nuclei puncta with GFP signal per worm as shown in A. n ≥ 15 worms. (C) Representative photomicrographs of the ADF neurotransmitter exocytosis reporter (ythEX766[srh-142p-SYP-Phluorin+unc-119(+)]) in control and srh-142p::TeTx::SL2::mCherry expressing animals. For the control group, mCherry channel image was not acquired (N/A) due to the absence of mCherry expression. The imaging used Z-stacks. Scale bar, 10 μm. (D) Representative photomicrographs of hsp-6 reporter (zcIs3[hsp-6p::gfp]) expression in animals treated with vehicle control or 50 mM serotonin (5-HT). (E) Quantification of hsp-6p::gfp expression. The genotypes are as in D. n ≥ 15 worms. (F) Representative photomicrographs of hsp-6 reporter expression in rgef-1p::Q40::yfp; tmbim-2(yth26) animals treated with vehicle control or 50 mM 5-HT. (G) Quantification of hsp-6p::gfp expression. The genotypes are as in F. n ≥ 15 worms. (H) Representative TEM images of synaptic profiles, including WT, tmbim-2(yth57), and tmbim-2(yth26). Arrows indicate active zone; Scale bar, 200 nm. (I) Quantification of the number of synaptic vesicles at the active zone as shown in H. n ≥ 5 worms. (J) Quantification of the size of synaptic vesicles at the active zone as shown in H. n ≥ 5 worms. (K) Dot plot shows the average expression of Tmbim2 in 17 mouse organs or tissue at the age of 3 mo. WBC, white blood cells; BAT, brown adipose tissue; SCAT, subcutaneous adipose tissue; GAT, gonadal adipose tissue; MAT, mesenteric adipose tissue. (L) Representative confocal photomicrograph of animals spermatheca expressing TMBIM-2 at different ages. The imaging used z-planes. Scale bar, 20 μm. (M) Quantitative analysis of TMBIM-2::GFP protein levels in ROI shown in L. n ≥ 8 worms. (N) Survival analysis of WT (blue) and tmbim-2 (yth26) (yellow); n ≥ 80 worms. (O) Survival analysis of control (blue) and ADF neuron overexpressing TMBIM-2 (yellow); n ≥ 80 worms. ***P < 0.001, **P < 0.01, ns denotes P > 0.05 via unpaired two-tailed Student’s t test. In N and O, we used The Log-rank (Mantel-Cox) test for statistical analysis. Error bars, SEM.
