Figure 8. Overexpression of human primary miR-125b-1 in the adult fat tissue promotes longevity and increases TAG levels under both AL and DR conditions.

(A) Schematic of Drosophila melanogaster primary miR-125 (pri miR-125) transcript and the Homo sapiens primary miR-125b-1 (pri hsa miR-125b-1) indicating that the processed miRNA sequence in fruit flies and humans is identical. (B) Quantitative RT-PCR of miR-125 from abdominal tissue of FB GS >UAS chinmo flies in presence of RU-486 (bars with red and blue pattern) or in absence of RU-486 (bars with solid red and blue color) in flies that were fed Ad libitum (AL) (red) or DR diet (blue) for 20 days. Expression levels were normalized to SnoR442. Values are mean ± SD, n = 6. (C) Overexpression of hsa pri miR-125 increases lifespan in flies that are fed an AL or DR diet (Compare red and blue dotted lines with red and blue solid line) (See Figure 8—source data 1A–B for number of flies and median and maximum lifespan). The p values and Χ² values indicate that the increase in lifespan is greater under DR conditions. (D) FB GS >UAS hsa pri miR-125b-1 flies store increased levels of triglycerides (TAG) when fed an AL or DR diet. Quantitation of TAG stored levels in -RU-486 and +RU-486 fed 20 d old FBGS >UAS hsa pri miR-125b-1 flies. (E) Fat bodies/abdomens of female flies were dissected and stained for the content and diameter of lipid droplet (LD) (red are lipid droplets stained with Nile red and blue is Dapi). Scale bar, 25 μm. (F) Quantitation of lipid droplet (LD) diameter in (E). Quantitation of 15 largest LDs in five samples per condition. Error bars represent ± SD. (G, H) Overexpression of hsa pri miR-125 upregulates fasn1 but not fatp under AL conditions. RT-PCR quantitation of mRNA levels of fasn1 (G) and fatp (H) in the fat tissue of FB GS >UAS hsa pri mir-125 flies that were fed an AL (red and pink bars) or DR diet (dark blue and light blue) under uninduced (red and dark blue bars) and induced (pink and light blue bars) conditions for 20 days. Expression levels were normalized to Actin5c. Values are mean ± SD, n ≥ 6. Genotypes of strains used in this figure: (B-H) FBGS/+; UAS has pri miR-125b-1: w¹¹¹⁸; P{w[+mW.hs]=Switch1}106/+; P{v+, UAS-hsa miR-125b-1} VK00033/+.

Figure 8—figure supplement 1. Effect of RU on lifespan of UAS pri hsa miR-125b-1>w¹¹¹⁸ flies.

To test if RU486 affects lifespan of flies expressing the UAS pri hsa miR-125b-1 transgene. Median lifespan in: AL- is 30 days; AL + is 28 days: DR- is 52 days and DR +is 46 days. Please refer to Figure 8—figure supplement 1—source data 1 for the second replicate of this experiment. Genotype of strain used in this Figure: +/+; UAS pri has miR-125b-1: w¹¹¹⁸; +/+; P{v+, UAS-hsa miR-125b-1} VK00033/+.