Amino acids activate mTOR complex 1 via Ca2+/CaM signaling to hVps34

Excess levels of circulating amino acids (AAs) play a causal role in specific human pathologies, including obesity and type 2 diabetes. Moreover, obesity and diabetes are contributing factors in the development of cancer, with recent studies suggesting that this link is mediated in part by AA activation of mammalian target of rapamycin (mTOR) Complex 1. AAs appear to mediate this response through class III phosphatidylinositol 3-kinase (PI3K), or human vacuolar protein sorting 34 (hVps34), rather than through the canonical class I PI3K pathway used by growth factors and hormones. Here we show that AAs induce a rise in intracellular Ca²⁺ ([Ca²⁺]_i), which triggers mTOR Complex 1 and hVps34 activation. We demonstrate that the rise in [Ca²⁺]_i increases the direct binding of Ca²⁺/calmodulin (CaM) to an evolutionarily conserved motif in hVps34 that is required for lipid kinase activity and increased mTOR Complex 1 signaling. These findings have important implications regarding the basic signaling mechanisms linking metabolic disorders with cancer progression.     

Phylogenesis of Brain Glutamic Acid Decarboxylase from Vertebrates: Immunochemical Studies

Brain high-speed supernatants from various lower and higher vertebrates were subjected to sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, electroblot on nitrocellulose membranes, and immunolabelling using an anti-glutamic acid decarboxylase (anti-GAD) antiserum prepared from rat antigen. Rat brain extracts showed two distinct immunolabelled bands (MW 59,000 and 62,000 daltons). The molecular weight of the native enzyme was 120,000 daltons. The immunoblot pattern was not affected by a 3-h incubation of the homogenate. In the substantia nigra, the decrease in the immunolabelling of both bands corresponded very closely to the decrease of GAD activity following lesioning of the striato-nigral pathway. Moreover, experiments with preadsorbed antiserum showed that both subunits have common antigenic determinants. The immunolabelling was consistently more intense over the lightest band. The autoradiography of immunoprecipitated rat brain GAD, iodinated prior to electrophoresis, revealed two radiolabelled bands corresponding to the two immunolabelled ones. Their radioactivity was found in a one-to-five ratio which closely paralleled their respective immunolabelling intensity. Thus, the two subunits recognized by the antiserum are not present in stoichiometric proportions in the rat brain high-speed supernatant. These findings suggest the existence of two homodimeric GAD with common antigenic determinants which are present in different amounts. Immunoprecipitation curves of brain GAD from rat, mouse, rabbit, monkey, human, quail, frog, and trout were similar, with a less than 10-fold maximum shift in affinity for GAD. GAD immunoblots from the various higher vertebrates showed a pattern similar to that obtained in rat.(ABSTRACT TRUNCATED AT 250 WORDS)     

Bothrops lanceolatus snake venom impairs mitochondrial respiration and induces DNA release in human heart preparation

IntroductionEnvenomations by Bothrops snakebites can induce overwhelming systemic inflammation ultimately leading to multiple organ system failure and death. Release of damage-associated molecular pattern molecules (DAMPs), in particular of mitochondrial origin, has been implicated in the pathophysiology of the deregulated innate immune response.ObjectiveTo test whether whole Bothrops lanceolatus venom would induce mitochondrial dysfunction and DAMPs release in human heart preparations.MethodsHuman atrial trabeculae were obtained during cannulation for cardiopulmonary bypass from patients who were undergoing routine coronary artery bypass surgery. Cardiac fibers were incubated with vehicle and whole Bothrops lanceolatus venom for 24hr before high-resolution respirometry, mitochondrial membrane permeability evaluation and quantification of mitochondrial DNA.ResultsCompared with vehicle, incubation of human cardiac muscle with whole Bothrops lanceolatus venom for 24hr impaired respiratory control ratio and mitochondrial membrane permeability. Levels of mitochondrial DNA increased in the medium of cardiac cell preparation incubated with venom of Bothrops lanceolatus.ConclusionOur study suggests that whole venom of Bothrops lanceolatus impairs mitochondrial oxidative phosphorylation capacity and increases mitochondrial membrane permeability. Cardiac mitochondrial dysfunction associated with mitochondrial DAMPs release may alter myocardium function and engage the innate immune response, which may both participate to the cardiotoxicity occurring in patients with severe envenomation.     

Maternal transmission of SIVsmm in rhesus macaques

Fifteen SIV-infected rhesus monkeys delivered 13 livebirths and two stillbirths; one livebirth died at three days of age. While all infants were culture-negative for SIV at birth, nine had maternal antibodies that disappeared by six months of age. Three infants subsequently seroconverted and became virus positive at 9–15 months. Milk samples from all mothers were virus-negative at parturition but samples from four animals were virus-positive at nine and 12 months. This study documents maternal transmission of SIV and suggests transmission by breast-feeding.     

Patterns of Gene Expression in a Scleractinian Coral Undergoing Natural Bleaching

Coral bleaching is a major threat to coral reefs worldwide and is predicted to intensify with increasing global temperature. This study represents the first investigation of gene expression in an Indo-Pacific coral species undergoing natural bleaching which involved the loss of algal symbionts. Quantitative real-time polymerase chain reaction experiments were conducted to select and evaluate coral internal control genes (ICGs), and to investigate selected coral genes of interest (GOIs) for changes in gene expression in nine colonies of the scleractinian coral Acropora millepora undergoing bleaching at Magnetic Island, Great Barrier Reef, Australia. Among the six ICGs tested, glyceraldehyde 3-phosphate dehydrogenase and the ribosomal protein genes S7 and L9 exhibited the most constant expression levels between samples from healthy-looking colonies and samples from the same colonies when severely bleached a year later. These ICGs were therefore utilised for normalisation of expression data for seven selected GOIs. Of the seven GOIs, homologues of catalase, C-type lectin and chromoprotein genes were significantly up-regulated as a result of bleaching by factors of 1.81, 1.46 and 1.61 (linear mixed models analysis of variance, P < 0.05), respectively. We present these genes as potential coral bleaching response genes. In contrast, three genes, including one putative ICG, showed highly variable levels of expression between coral colonies. Potential variation in microhabitat, gene function unrelated to the stress response and individualised stress responses may influence such differences between colonies and need to be better understood when designing and interpreting future studies of gene expression in natural coral populations.     

A set of accessible enhancers enables the initial response of breast cancer cells to physiological progestin concentrations

Here, we report that in T47D breast cancer cells 50 pM progestin is sufficient to activate cell cycle entry and the progesterone gene expression program. At this concentration, equivalent to the progesterone blood levels found around the menopause, progesterone receptor (PR) binds only to 2800 genomic sites, which are accessible to ATAC cleavage prior to hormone exposure. These highly accessible sites (HAs) are surrounded by well-organized nucleosomes and exhibit breast enhancer features, including estrogen receptor alpha (ERα), higher FOXA1 and BRD4 (bromodomain containing 4) occupancy. Although HAs are enriched in RAD21 and CTCF, PR binding is the driving force for the most robust interactions with hormone-regulated genes. HAs show higher frequency of 3D contacts among themselves than with other PR binding sites, indicating colocalization in similar compartments. Gene regulation via HAs is independent of classical coregulators and ATP-activated remodelers, relying mainly on MAP kinase activation that enables PR nuclear engagement. HAs are also preferentially occupied by PR and ERα in breast cancer xenografts derived from MCF-7 cells as well as from patients, indicating their potential usefulness as targets for therapeutic intervention.     

A general and efficient method for estimating continuous IBD functions for use in genome scans for QTL

Background  

Identity by descent (IBD) matrix estimation is a central component in mapping of Quantitative Trait Loci (QTL) using variance component models. A large number of algorithms have been developed for estimation of IBD between individuals in populations at discrete locations in the genome for use in genome scans to detect QTL affecting various traits of interest in experimental animal, human and agricultural pedigrees. Here, we propose a new approach to estimate IBD as continuous functions rather than as discrete values.