Biology2015, 4(1), 200-215; doi:10.3390/biology4010200 (registering DOI) - published 4 March 2015 Show/Hide Abstract
Abstract: The steroidogenic acute regulatory protein (STAR) protein expression is required for cholesterol transport into mitochondria to initiate steroidogenesis in the adrenal and gonads. STAR is synthesized as a 37 kDa precursor protein which is targeted to the mitochondria and imported and processed to an intra-mitochondrial 30 kDa protein. Tropic hormone stimulation of the cAMP-dependent protein kinase A (PKA) signaling pathway is the major contributor to the transcriptional and post-transcriptional regulation of STAR synthesis. Many studies have focused on the mechanisms of cAMP-PKA mediated control of STAR synthesis while there are few reports on STAR degradation pathways. The objective of this study was to determine the effect of cAMP-PKA-dependent signaling on STAR protein stability. We have used the cAMP-PKA responsive Y1 mouse adrenocortical cells and the PKA-deficient Kin-8 cells to measure STAR phosphorylation and protein half-life. Western blot analysis and standard radiolabeled pulse-chase experiments were used to determine STAR phosphorylation status and protein half-life, respectively. Our data demonstrate that PKA-dependent STAR phosphorylation does not contribute to 30 kDa STAR protein stability in the mitochondria. We further show that inhibition of the 26S proteasome does not block precursor STAR phosphorylation or steroid production in Y1 cells. These data suggest STAR can maintain function and promote steroidogenesis under conditions of proteasome inhibition.
Biology2015, 4(1), 187-199; doi:10.3390/biology4010187 (registering DOI) - published 4 March 2015 Show/Hide Abstract
Abstract: The reason why metabolic rate often scales allometrically (disproportionately) with body mass has been debated for decades. A critical question concerns whether metabolic scaling is controlled intrinsically at the intracellular level or systemically at the organismal level. Recently, the relative importance of these effects has been tested by examining the metabolic rates of cultured dermal fibroblast and skeletal muscle cells in relation to donor body mass of a variety of birds and mammals. The lack of a relationship between in vitro cellular metabolic rates and body mass suggests that systemic effects, not intrinsic cellular effects are responsible for allometric metabolic scaling observed in whole organisms. Influential resource-transport network theory claims that the most important systemic effect involved is body-size related resource-supply limits to metabolizing cells. However, comparisons of in vitro cellular metabolic rates with scaling relationships for in vivo (basal) metabolic rates suggest that other systemic effects, such as body-size dependent biological regulation and tissue composition may also have major, perhaps more important effects. Furthermore, systemic effects must ultimately act at the cellular level, for example, by induced variation in the function, structure and intracellular densities of mitochondria. The mechanistic pathways involved require further study.
Abstract: Anodal transcranial Direct Current Stimulation (tDCS) has been shown to be an effective non-invasive brain stimulation method for improving cognitive and motor functioning in patients with neurological deficits. tDCS over motor cortex (M1), for instance, facilitates motor learning in stroke patients. However, the literature on anodal tDCS effects on motor learning in healthy participants is inconclusive, and the effects of tDCS on visuo-motor integration are not well understood. In the present study we examined whether tDCS over the contralateral motor cortex enhances learning of grip-force output in a visually guided feedback task in young and neurologically healthy volunteers. Twenty minutes of 1 mA anodal tDCS were applied over the primary motor cortex (M1) contralateral to the dominant (right) hand, during the first half of a 40 min power-grip task. This task required the control of a visual signal by modulating the strength of the power-grip for six seconds per trial. Each participant completed a two-session sham-controlled crossover protocol. The stimulation conditions were counterbalanced across participants and the sessions were one week apart. Performance measures comprised time-on-target and target-deviation, and were calculated for the periods of stimulation (or sham) and during the afterphase respectively. Statistical analyses revealed significant performance improvements over the stimulation and the afterphase, but this learning effect was not modulated by tDCS condition. This suggests that the form of visuomotor learning taking place in the present task was not sensitive to neurostimulation. These null effects, together with similar reports for other types of motor tasks, lead to the proposition that tDCS facilitation of motor learning might be restricted to cases or situations where the motor system is challenged, such as motor deficits, advanced age, or very high task demand.
Abstract: Constrained peptides are an attractive class as affinity reagents or drug leads owing to their excellent binding properties. Many kinds of these peptides, such as cyclic peptides containing disulfide bridges, are found in nature or designed artificially by directed evolution. However, confirming the binding properties of the disulfide-rich peptides can be generally difficult, because of oxidative folding problems in the preparation steps. Therefore, a method for evaluating the binding properties of such peptides rapidly and easily is required. Here, we report an easy and rapid method for preparing biotin-attached peptides containing disulfide bridges or a chemical cross-linker using a cell-free translation system and a puromycin-linker, which is applicable to pull-down assays for protein (or peptide) molecular interaction analysis.
Abstract: Oncostatin M (OSM) is a cytokine belonging to the gp130 family, whose members serve pleiotropic functions. However, several actions of OSM are unique from those of other gp130 cytokines, and these actions may have critical roles in inflammatory mechanisms influencing several metabolic and biological functions of insulin-sensitive tissues. In this review, the actions of OSM in adipose tissue and liver are discussed, with an emphasis on lipid metabolism.
Abstract: In eukaryotic cells, mitochondria host ancient essential bioenergetic and biosynthetic pathways. LYR (leucine/tyrosine/arginine) motif proteins (LYRMs) of the Complex1_LYR-like superfamily interact with protein complexes of bacterial origin. Many LYR proteins function as extra subunits (LYRM3 and LYRM6) or novel assembly factors (LYRM7, LYRM8, ACN9 and FMC1) of the oxidative phosphorylation (OXPHOS) core complexes. Structural insights into complex I accessory subunits LYRM6 and LYRM3 have been provided by analyses of EM and X-ray structures of complex I from bovine and the yeast Yarrowia lipolytica, respectively. Combined structural and biochemical studies revealed that LYRM6 resides at the matrix arm close to the ubiquinone reduction site. For LYRM3, a position at the distal proton-pumping membrane arm facing the matrix space is suggested. Both LYRMs are supposed to anchor an acyl-carrier protein (ACPM) independently to complex I. The function of this duplicated protein interaction of ACPM with respiratory complex I is still unknown. Analysis of protein-protein interaction screens, genetic analyses and predicted multi-domain LYRMs offer further clues on an interaction network and adaptor-like function of LYR proteins in mitochondria.