Although selenophosphate synthetase 1 (SPS1/SelD) is an essential gene in Drosophila, its function has not been determined. To elucidate its intracellular role, we targeted the removal of SPS1/SelD mRNA in Drosophila SL2 cells using RNA interference technology that led to the formation of vacuole-like globular structures. Surprisingly, these structures were identified as megamitochondria, and only depolarized mitochondria developed into megamitochondria. The mRNA levels of l(2)01810 and glutamine synthetase 1 (GS1) were increased by SPS1/SelD knockdown. Blocking the expression of GS1 and l(2)01810 completely inhibited the formation of megamitochondria induced by loss of SPS1/SelD activity and decreased the intracellular levels of glutamine to those of control cells suggesting that the elevated level of glutamine is responsible for megamitochondrial formation. Overexpression of GS1 and l(2)01810 had a synergistic effect on the induction of megamitochondrial formation and on the synthesis of glutamine suggesting that l(2)01810 is involved in glutamine synthesis presumably by activating GS1. Our results indicate that, in Drosophila, SPS1/SelD regulates the intracellular glutamine by inhibiting GS1 and l(2)01810 expression and that elevated levels of glutamine lead to a nutritional stress that provides a signal for megamitochondrial formation.Selenium is an essential trace element in the diet of humans and many other life forms. It provides many health benefits such as roles in preventing cancer and heart disease, serving as an antiviral agent, stimulating the immune system, reactive oxygen species (ROS) 4 scavenging, and male reproduction (1-6). Many of the benefits of selenium are most likely due to the presence of this element in selenoproteins as the amino acid selenocysteine (Sec) (7-9). Sec is the 21st amino acid in the genetic code (10 -12) and is incorporated into selenoproteins in response to UGA Sec codons (13,14). The active donor of selenium in Sec biosynthesis is monoselenophosphate (15), which is synthesized from selenite and ATP by an enzyme designated as selenophosphate synthetase (SPS) (16). There are two isoforms of SPS in higher eukaryotes, SPS1/SelD and SPS2, whereas only one type of SPS (SelD) exists in lower eukaryotes and eubacteria (17). The sequences of SPS1/SelD and SPS2 are highly conserved. For example, the amino acid sequence homology between human SPS1/SelD and SPS2 is 72% and that between Drosophila SPS1/SelD and SPS2 is ϳ45%. One of the major differences between SPS1/SelD and SPS2 is that SPS1/ SelD has an arginine at the position corresponding to Sec in SPS2 (18,19).Initially, both SPS1/SelD and SPS2 were thought to be involved in selenophosphate synthesis. However, it was subsequently shown that only SPS2 catalyzes selenophosphate synthesis. In in vitro experiments, SPS2 synthesized selenophosphate from selenide and ATP, but SPS1/SelD did not have this activity (20). Knockdown of SPS2 in NIH3T3 cells led to the loss of selenoprotein biosynthesis, whereas the inhibition of SPS1/ SelD...

Background/Aims: Stress is closely related to the deterioration of digestive disease. Melatonin has potent anti-inflammatory properties. The objective of this study was to determine the effect of water stress (WS) and sleep deprivation (SD) on intestinal microbiota and roles of melatonin in stressful condition. Methods: We used C57BL/6 mice and specially designed water bath for stress and SD for 10 days. We measured melatonin concentrations in serum, feces, and colon tissues by high-performance liquid chromatography. Genomic DNA was extracted from feces and amplified using primers targeting V3 to V4 regions of bacterial 16S ribosomal RNA genes. Results: Compared to the control, melatonin concentration was lower in the WS and SD. Fecal concentration was 0.132 pg/mL in control, 0.062 pg/mL in WS, and 0.068 pg/mL in SD. In colon tissue, it was 0.45 pg/mL in control, 0.007 pg/mL in WS, and 0.03 pg/mL in SD. After melatonin treatment, melatonin concentrations in feces and colon tissue were recovered to the level of control. Metagenomic analysis of microbiota showed abundance in colitogenic microbiota in WS and SD. Melatonin injection attenuated this harmful effect. WS and SD showed decreased <i>Lactobacillales</i> and increased <i>Erysipelotrichales</i> and <i>Enterobacteriales</i>. Melatonin treatment increased <i>Akkermansia muciniphila</i> and <i>Lactobacillus</i> and decreased <i>Bacteroides massiliensis</i> and <i>Erysipelotrichaceae</i>. Conclusions: This study showed that stress and SD could affect intestinal dysbiosis and increase colitogenic microbiota, which could contribute to the aggravating digestive disease. Melatonin concentrations in feces and colon tissue decreased under WS and SD. Melatonin treatment brought recovery of melatonin concentration in colon tissue and modulating dysbiosis of intestinal microbiota.

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