Evidence for direct roles of two additional factors, SECp43 and soluble liver antigen, in the selenoprotein synthesis machinery

Selenocysteine (Sec) is inserted into selenoproteins co-translationally with the help of various cis- and trans-acting factors. The specific mechanisms of Sec biosynthesis and insertion into protein in eukaryotic cells, however, are not known. Two proteins, SECp43 and the soluble liver antigen (SLA), were previously reported to interact with tRNA([Ser]Sec), but their functions remained elusive. Herein, we report that knockdown of SECp43 in NIH3T3 or TCMK-1 cells using RNA interference technology resulted in a reduction in the level of methylation at the 2'-hydroxylribosyl moiety in the wobble position (Um34) of Sec tRNA([Ser]Sec), and consequently reduced glutathione peroxidase 1 expression. Double knockdown of SECp43 and SLA resulted in decreased selenoprotein expression. SECp43 formed a complex with Sec tRNA([Ser]Sec) and SLA, and the targeted removal of one of these proteins affected the binding of the other to Sec tRNA([Ser]Sec). SECp43 was located primarily in the nucleus, whereas SLA was found in the cytoplasm. Co-transfection of both proteins resulted in the nuclear translocation of SLA suggesting that SECp43 may also promote shuttling of SLA and Sec tRNA([Ser]Sec) between different cellular compartments. Taken together, these data establish the role of SECp43 and SLA in selenoprotein biosynthesis through interaction with tRNA([Ser]Sec) in a multiprotein complex. The data also reveal a role of SECp43 in regulation of selenoprotein expression by affecting the synthesis of Um34 on tRNA([Ser]Sec) and the intracellular location of SLA.     

Thioredoxin reductase 1 deficiency reverses tumor phenotype and tumorigenicity of lung carcinoma cells

Dietary selenium has potent cancer prevention activity. Both low molecular weight selenocompounds and selenoproteins are implicated in this effect. Thioredoxin reductase 1 (TR1) is one of the major antioxidant and redox regulators in mammals that supports p53 function and other tumor suppressor activities. However, this selenium-containing oxidoreductase is also overexpressed in many malignant cells and has been proposed as a target for cancer therapy. To further assess the role of TR1 in the malignancy process, we used RNA interference technology to decrease its expression in mouse lung carcinoma (LLC1) cells. Stable transfection of LLC1 cells with a small interfering RNA construct that specifically targets TR1 removal manifested a reversal in the morphology and anchorage-independent growth properties of these cancer cells that made them similar to those of normal cells. The expression of at least two cancer-related protein mRNAs, Hgf and Opn1, were reduced dramatically in the TR1 knockdown cells. Mice injected with the TR1 knockdown showed a dramatic reduction in tumor progression and metastasis compared with those mice injected with the corresponding control vector. In addition, tumors that arose from injected TR1 knockdown cells lost the targeting construct, suggesting that TR1 is essential for tumor growth in mice. These observations provide direct evidence that the reduction of TR1 levels in malignant cells is antitumorigenic and suggest that the enzyme is a prime target for cancer therapy.     

Redox regulation of cell signaling by selenocysteine in mammalian thioredoxin reductases.

The intracellular generation of reactive oxygen species, together with the thioredoxin and glutathione systems, is thought to participate in redox signaling in mammalian cells. The activity of thioredoxin is dependent on the redox status of thioredoxin reductase (TR), the activity of which in turn is dependent on a selenocysteine residue. Two mammalian TR isozymes (TR2 and TR3), in addition to that previously characterized (TR1), have now been identified in humans and mice. All three TR isozymes contain a selenocysteine residue that is located in the penultimate position at the carboxyl terminus and which is encoded by a UGA codon. The generation of reactive oxygen species in a human carcinoma cell line was shown to result in both the oxidation of the selenocysteine in TR1 and a subsequent increase in the expression of this enzyme. These observations identify the carboxyl-terminal selenocysteine of TR1 as a cellular redox sensor and support an essential role for mammalian TR isozymes in redox-regulated cell signaling.     

Inter-species variation in yolk steroid levels and a cowbird-host comparison

We examined variability in yolk hormone levels among songbird species and the role of yolk steroids as a mechanism for enhanced exploitation of hosts by the parasitic brown-headed cowbird Molothrus ater . Within-clutch variation in yolk steroids has been found in several avian species in single species studies, but few comparisons have been made among species. We found a large range of differences in yolk testosterone among the seven passerine species examined, with significant differences between those at the high end (song sparrow Melospiza melodia , red-winged blackbird Agelaius phoeniceus , and house sparrow, Passer domesticus ) and those at the low end (eastern phoebe Sayornis phoebe , and house finch Carpodacus mexicanus ). We also found that the testosterone level in cowbird eggs was intermediate in relation to host species levels and was significantly lower than that in three common cowbird hosts (song sparrow, red-winged blackbird, and house sparrow), but not significantly different from three others. Geographical comparisons of yolk testosterone levels in all cowbird subspecies and populations from several regions showed no significant differences, though a trend that deserves further exploration was the pattern of lowest level in the ancestral population of cowbirds in the central prairies and of highest level in the northwestern population where range invasion occurred approximately 40 years ago. The levels of 17 beta-estradiol were similar in the seven songbird species examined, which is consistent with current hypotheses that this hormone plays a role in embryonic sexual differentiation. Further investigation is needed to determine whether the large differences observed among species in absolute level of yolk testosterone are the relevant focal point or whether target tissue sensitivity differences mediate the effects of this yolk steroid, particularly between parasitic and non-parasitic species.     

Effects of L-carnitine L-tartrate supplementation on muscle oxygenation responses to resistance exercise

Previous research has shown that L-carnitine L-tartrate (LCLT) supplementation beneficially affects markers of hypoxic stress following resistance exercise. However, the mechanism of this response is unclear. Therefore, the primary purpose of this study was to determine the effects of LCLT supplementation on muscle tissue oxygenation during and after multiple sets of squat exercise. Nine healthy, previously resistance-trained men (25.2 +/- 6.years, 91.2 +/- 10.2 kg, 180.2 +/- 6.3 cm) ingested 2 g.d of LCLT or an identical placebo for 23 days in a randomized, balanced, crossover, double-blind, placebo-controlled, repeated-measures study design. On day 21, forearm muscle oxygenation was measured during and after an upper arm occlusion protocol using near infrared spectroscopy (NIRS), which measures the balance of oxygen delivery in relation to oxygen consumption. On day 22, subjects performed 5 sets of 15 to 20 repetitions of squat exercise with corresponding measures of thigh muscle oxygenation, via NIRS, and serial blood draws. Compared to the placebo trial, muscle oxygenation was reduced in the LCLT trial during upper arm occlusion and following each set of resistance exercise. Despite reduced oxygenation, plasma malondealdehyde, a marker of membrane damage, was attenuated during the LCLT trial. There were no differences between trials in the vasoactive substance prostacyclin. In conclusion, because oxygen delivery was occluded during the forearm protocol, it is proposed that enhanced oxygen consumption mediated the reduced muscle oxygenation during the LCLT trial. Enhanced oxygen consumption would explain why hypoxic stress was attenuated with LCLT supplementation.     

Comparative analysis of selenocysteine machinery and selenoproteome gene expression in mouse brain identifies neurons as key functional sites of selenium in mammals

Although dietary selenium (Se) deficiency results in phenotypes associated with selenoprotein depletion in various organs, the brain is protected from Se loss. To address the basis for the critical role of Se in brain function, we carried out comparative gene expression analyses for the complete selenoproteome and associated biosynthetic factors. Using the Allen Brain Atlas, we evaluated 159 regions of adult mouse brain and provided experimental analyses of selected selenoproteins. All 24 selenoprotein mRNAs were expressed in the mouse brain. Most strikingly, neurons in olfactory bulb, hippocampus, cerebral cortex, and cerebellar cortex were exceptionally rich in selenoprotein gene expression, in particular in GPx4, SelK, SelM, SelW, and Sep15. Over half of the selenoprotein genes were also expressed in the choroid plexus. A unique expression pattern was observed for one of the highly expressed selenoprotein genes, SelP, which we suggest to provide neurons with Se. Cluster analysis of the expression data linked certain selenoproteins and selenocysteine machinery genes and suggested functional linkages among selenoproteins, such as that between SelM and Sep15. Overall, this study suggests that the main functions of selenium in mammals are confined to certain neurons in the brain.     

Effects of stretching on upper-body muscular performance

The purpose of this investigation was to examine the influence of upper-body static stretching and dynamic stretching on upper-body muscular performance. Eleven healthy men, who were National Collegiate Athletic Association Division I track and field athletes (age, 19.6 +/- 1.7 years; body mass, 93.7 +/- 13.8 kg; height, 183.6 +/- 4.6 cm; bench press 1 repetition maximum [1RM], 106.2 +/- 23.0 kg), participated in this study. Over 4 sessions, subjects participated in 4 different stretching protocols (i.e., no stretching, static stretching, dynamic stretching, and combined static and dynamic stretching) in a balanced randomized order followed by 4 tests: 30% of 1 RM bench throw, isometric bench press, overhead medicine ball throw, and lateral medicine ball throw. Depending on the exercise, test peak power (Pmax), peak force (Fmax), peak acceleration (Amax), peak velocity (Vmax), and peak displacement (Dmax) were measured. There were no differences among stretch trials for Pmax, Fmax, Amax, Vmax, or Dmax for the bench throw or for Fmax for the isometric bench press. For the overhead medicine ball throw, there were no differences among stretch trials for Vmax or Dmax. For the lateral medicine ball throw, there was no difference in Vmax among stretch trials; however, Dmax was significantly larger (p 相似文献