Multivariable association discovery in population-scale meta-omics studies

It is challenging to associate features such as human health outcomes, diet, environmental conditions, or other metadata to microbial community measurements, due in part to their quantitative properties. Microbiome multi-omics are typically noisy, sparse (zero-inflated), high-dimensional, extremely non-normal, and often in the form of count or compositional measurements. Here we introduce an optimized combination of novel and established methodology to assess multivariable association of microbial community features with complex metadata in population-scale observational studies. Our approach, MaAsLin 2 (Microbiome Multivariable Associations with Linear Models), uses generalized linear and mixed models to accommodate a wide variety of modern epidemiological studies, including cross-sectional and longitudinal designs, as well as a variety of data types (e.g., counts and relative abundances) with or without covariates and repeated measurements. To construct this method, we conducted a large-scale evaluation of a broad range of scenarios under which straightforward identification of meta-omics associations can be challenging. These simulation studies reveal that MaAsLin 2’s linear model preserves statistical power in the presence of repeated measures and multiple covariates, while accounting for the nuances of meta-omics features and controlling false discovery. We also applied MaAsLin 2 to a microbial multi-omics dataset from the Integrative Human Microbiome (HMP2) project which, in addition to reproducing established results, revealed a unique, integrated landscape of inflammatory bowel diseases (IBD) across multiple time points and omics profiles.     

Loss of prion protein control of glucose metabolism promotes neurodegeneration in model of prion diseases

Corruption of cellular prion protein (PrP^C) function(s) at the plasma membrane of neurons is at the root of prion diseases, such as Creutzfeldt-Jakob disease and its variant in humans, and Bovine Spongiform Encephalopathies, better known as mad cow disease, in cattle. The roles exerted by PrP^C, however, remain poorly elucidated. With the perspective to grasp the molecular pathways of neurodegeneration occurring in prion diseases, and to identify therapeutic targets, achieving a better understanding of PrP^C roles is a priority. Based on global approaches that compare the proteome and metabolome of the PrP^C expressing 1C11 neuronal stem cell line to those of PrP^null-1C11 cells stably repressed for PrP^C expression, we here unravel that PrP^C contributes to the regulation of the energetic metabolism by orienting cells towards mitochondrial oxidative degradation of glucose. Through its coupling to cAMP/protein kinase A signaling, PrP^C tones down the expression of the pyruvate dehydrogenase kinase 4 (PDK4). Such an event favors the transfer of pyruvate into mitochondria and its conversion into acetyl-CoA by the pyruvate dehydrogenase complex and, thereby, limits fatty acids β-oxidation and subsequent onset of oxidative stress conditions. The corruption of PrP^C metabolic role by pathogenic prions PrP^Sc causes in the mouse hippocampus an imbalance between glucose oxidative degradation and fatty acids β-oxidation in a PDK4-dependent manner. The inhibition of PDK4 extends the survival of prion-infected mice, supporting that PrP^Sc-induced deregulation of PDK4 activity and subsequent metabolic derangements contribute to prion diseases. Our study posits PDK4 as a potential therapeutic target to fight against prion diseases.     

Temporal metabonomic modeling of l-arginine-induced exocrine pancreatitis

The time-related metabolic responses to l-arginine (ARG)-induced exocrine pancreatic toxicity were investigated using single ip doses of 1,000 and 4,000 mg/kg body weight over a 7 day experimental period in male Sprague-Dawley rats. Sequential timed urine and plasma samples were analyzed using high resolution (1)H NMR spectroscopy together with complementary clinical chemistry and histopathology analyses. Principal components analysis (PCA) and orthogonal projection on latent structures discriminant analysis (O-PLS-DA) were utilized to analyze the (1)H NMR data and to extract and identify candidate biomarkers and to construct metabolic trajectories post ARG administration. Low doses of ARG resulted in virtually no histopathological damage and distinct reversible metabolic response trajectories. High doses of ARG caused pancreatic acinar degeneration and necrosis and characteristic metabolic trajectory profiles with several distinct phases. The initial trajectory phase (0-8 h) involved changes in the urea cycle and transamination indicating a homeostatic response to detoxify excess ammonia generated from ARG catabolism. By 48 h, there was a notable enhancement of the excretion of the gut microbial metabolites, phenylacetylglycine (PAG), 4-cresol-glucuronide and 4-cresol-sulfate, suggesting that compromised pancreatic function impacts on the activity of the gut microbiota giving potential rise to a novel class of surrogate extragenomic biomarkers of pancreatic injury. The implied compromise of microbiotal function may also contribute to secondary hepatic and pancreatic toxic responses. We show here for the first time the value of metabonomic studies in investigating metabolic disruption due to experimental pancreatitis. The variety of observed systemic responses suggests that this approach may be of general value in the assessment of other animal models or human pancreatitis.     

Selective mitochondrial autophagy during erythroid maturation

Accumulating evidence suggests that autophagy can be selective in the clearance of organelles in yeast and in mammalian cells. We have observed that the sequestration of mitochondria by autophagosomes was defective in reticulocytes in the absence of Nix. Nix is required for the dissipation of mitochondrial membrane potential (DeltaPsim) during erythroid maturation. Moreover, pharmacological agents that induce the loss of DeltaPsim can restore the sequestration of mitochondria by autophagosomes and promote mitochondrial clearance in Nix(-/-) erythroid cells. Our data suggest that mitochondrial depolarization induces recognition and sequestration of mitochondria by autophagosomes. Elucidating the mechanisms underlying selective mitochondrial autophagy not only will help us to understand the mechanisms for erythroid maturation, but also may provide insights into mitochondrial quality control by autophagy in the protection against aging, cancer and neurodegenerative diseases.     

Minimising biomass/adsorbent interactions in expanded bed adsorption processes: a methodological design approach

Expanded bed adsorption (EBA) is an integrated technology for the primary recovery of proteins from crude feedstock. Interactions between solid matter in the feed suspension and fluidised adsorbent particles influence bed stability and therefore have a significant impact on protein adsorption in expanded beds. In order to design efficient and reliable EBA processes a strategy is needed, which allows to find operating conditions, where these adverse events do not take place. In this paper a methodological approach is presented, which allows systematic characterisation and minimisation of cell/adsorbent interactions with as little experimental effort as possible. Adsorption of BSA to the anion exchanger Streamline Q XL from a suspension containing S. cerevisiae cells was chosen as a model system with a strong affinity of the biomass towards the stationary phase. Finite bath biomass adsorption experiments were developed as an initial screening method to estimate a potential interference. The adhesiveness of S. cerevisiae to the anion exchanger could be reduced significantly by increasing the conductivity of the feedstock. A biomass pulse response method was used to find optimal operation conditions showing no cell/adsorbent interactions. A good correlation was found between the finite bath test and the pulse experiment for a variety of suspensions (intact yeast cells, E. coli homogenate and hybridoma cells) and adsorbents (Streamline Q XL, DEAE and SP), which allows to predict cell/adsorbent interactions in expanded beds just from finite bath adsorption tests. Under the optimised operating conditions obtained using the prior methods, the stability of the expanded bed was investigated during fluidisation in biomass containing feedstock (up to 15% yeast on wet weight basis) employing residence time distribution analysis and evaluation by an advanced model. Based on these studies threshold values were defined for the individual experiments, which have to be achieved in order to obtain an efficient EBA process. Breakthrough experiments were conducted to characterise the efficiency of BSA adsorption from S. cerevisiae suspensions in EBA mode under varying operating conditions. This allowed to correlate the stability of the expanded bed with its sorption efficiency and therefore could be used to verify the threshold values defined. The approach presented in this work provides a fast and simple way to minimise cell/adsorbent interactions and to define a window of operation for protein purification using EBA.     

The HA-2 minor histocompatibility antigen is derived from a diallelic gene encoding a novel human class I myosin protein.

Human minor histocompatibility Ags (mHag) present significant barriers to successful bone marrow transplantation. However, the structure of human mHag and the basis for antigenic disparities are still largely unknown. Here we report the identification of the gene encoding the human mHag HA-2 as a previously unknown member of the class I myosin family, which we have designated MYO1G. The gene is located on the short arm of chromosome 7. Expression of this gene is limited to cells of hemopoietic origin, in keeping with the previously defined tissue expression of the HA-2 Ag. RT-PCR amplification of MYO1G from different individuals led to the identification of two genetic variants, designated MYO1G(V) and MYO1G(M). The former encodes the peptide sequence previously shown to be the HA-2 epitope (YIGEVLVSV), whereas the latter shows a single amino acid change in this peptide (YIGEVLVSM). This change has only a modest effect on peptide binding to the class I MHC-restricted element HLA-A*0201, and a minimal impact on recognition by T cells when added exogenously to target cells. Nonetheless, as detected using either T cells or mass spectrometry, this amino acid change results in a failure of the latter peptide to be presented at the surface of cells that express MYO1G(M) endogenously. These studies have thus identified a new mHag-encoding gene, and thereby provide additional information about both the genetic origins of human mHag as well as the underlying basis of an Ag-positive vs Ag-negative state.     

Contents of 1,4-benzoxazin-3-ones and 2-benzoxazolinone from Stenandrium dulce (Nees)

Secondary metabolites, DIBOA, HBOA, 7-OH-HBOA and BOA, were isolated and quantified from S. dulce (Nees), a native species in Chile belonging to the Acanthaceae family. The highest DIBOA and HBOA contents were determined in leaves (9.25 mmol kg(-1) fr. wt) and root (6.81 mmol kg(-1) fr. wt), respectively. Aglycones, 7-OH-HBOA and HBOA, were isolated together from root extracts of Acanthaceae species. Both, HBOA and 7-OH-HBOA should be direct precursors in the biosynthesis of DIBOA and DIMBOA, respectively.