Reproductive performance of dairy cows from parturition to conception

International audienc

Transcriptional Profiling Uncovers a Network of Cholesterol-Responsive Atherosclerosis Target Genes

Despite the well-documented effects of plasma lipid lowering regimes halting atherosclerosis lesion development and reducing morbidity and mortality of coronary artery disease and stroke, the transcriptional response in the atherosclerotic lesion mediating these beneficial effects has not yet been carefully investigated. We performed transcriptional profiling at 10-week intervals in atherosclerosis-prone mice with human-like hypercholesterolemia and a genetic switch to lower plasma lipoproteins (Ldlr−/−Apo100/100 Mttpflox/flox Mx1-Cre). Atherosclerotic lesions progressed slowly at first, then expanded rapidly, and plateaued after advanced lesions formed. Analysis of lesion expression profiles indicated that accumulation of lipid-poor macrophages reached a point that led to the rapid expansion phase with accelerated foam-cell formation and inflammation, an interpretation supported by lesion histology. Genetic lowering of plasma cholesterol (e.g., lipoproteins) at this point all together prevented the formation of advanced plaques and parallel transcriptional profiling of the atherosclerotic arterial wall identified 37 cholesterol-responsive genes mediating this effect. Validation by siRNA-inhibition in macrophages incubated with acetylated-LDL revealed a network of eight cholesterol-responsive atherosclerosis genes regulating cholesterol-ester accumulation. Taken together, we have identified a network of atherosclerosis genes that in response to plasma cholesterol-lowering prevents the formation of advanced plaques. This network should be of interest for the development of novel atherosclerosis therapies

Alternative mRNA Editing in Trypanosomes Is Extensive and May Contribute to Mitochondrial Protein Diversity

The editing of trypanosome mitochondrial mRNAs produces transcripts necessary for mitochondrial functions including electron transport and oxidative phosphorylation. Precursor-mRNAs are often extensively edited by specific uridine insertion or deletion that is directed by small guide RNAs (gRNAs). Recently, it has been shown that cytochrome c oxidase subunit III (COXIII) mRNAs can be alternatively edited to encode a novel mitochondrial membrane protein composed of a unique hydrophilic N-terminal sequence of unknown function and the C-terminal hydrophobic segment of COXIII. To extend the analysis of alternative editing in Trypanosoma brucei we have constructed libraries with over 1100 full-length mitochondrial cDNAs and the sequences of over 1200 gRNA genes. Using this data, we show that alternative editing of COXIII, ATPase subunit 6 (A6), and NADH dehydrogenase subunits 7, 8 and 9 (ND7, 8, 9) mRNAs can produce novel open reading frames (ORFs). Several gRNAs potentially responsible for the alternative editing of these mRNAs were also identified. These findings show that alternative editing of mitochondrial mRNAs is common in T. brucei and expands the diversity of mitochondrial proteins in these organisms

Complete Mitochondrial Genome Sequence of Three Tetrahymena Species Reveals Mutation Hot Spots and Accelerated Nonsynonymous Substitutions in Ymf Genes

The ciliate Tetrahymena, a model organism, contains divergent mitochondrial (Mt) genome with unusual properties, where half of its 44 genes still remain without a definitive function. These genes could be categorized into two major groups of KPC (known protein coding) and Ymf (genes without an identified function). To gain insights into the mechanisms underlying gene divergence and molecular evolution of Tetrahymena (T.) Mt genomes, we sequenced three Mt genomes of T.paravorax, T.pigmentosa, and T.malaccensis. These genomes were aligned and the analyses were carried out using several programs that calculate distance, nucleotide substitution (dn/ds), and their rate ratios (ω) on individual codon sites and via a sliding window approach. Comparative genomic analysis indicated a conserved putative transcription control sequence, a GC box, in a region where presumably transcription and replication initiate. We also found distinct features in Mt genome of T.paravorax despite similar genome organization among these ∼47 kb long linear genomes. Another significant finding was the presence of at least one or more highly variable regions in Ymf genes where majority of substitutions were concentrated. These regions were mutation hotspots where elevated distances and the dn/ds ratios were primarily due to an increase in the number of nonsynonymous substitutions, suggesting relaxed selective constraint. However, in a few Ymf genes, accelerated rates of nonsynonymous substitutions may be due to positive selection. Similarly, on protein level the majority of amino acid replacements occurred in these regions. Ymf genes comprise half of the genes in Tetrahymena Mt genomes, so understanding why they have not been assigned definitive functions is an important aspect of molecular evolution. Importantly, nucleotide substitution types and rates suggest possible reasons for not being able to find homologues for Ymf genes. Additionally, comparative genomic analysis of complete Mt genomes is essential in identifying biologically significant motifs such as control regions

GABA-A Channel Subunit Expression in Human Glioma Correlates with Tumor Histology and Clinical Outcome

GABA (γ-aminobutyric acid) is the main inhibitory neurotransmitter in the CNS and is present in high concentrations in presynaptic terminals of neuronal cells. More recently, GABA has been ascribed a more widespread role in the control of cell proliferation during development where low concentrations of extrasynaptic GABA induce a tonic activation of GABA receptors. The GABA-A receptor consists of a ligand-gated chloride channel, formed by five subunits that are selected from 19 different subunit isoforms. The functional and pharmacological properties of the GABA-A channels are dictated by their subunit composition. Here we used qRT-PCR to compare mRNA levels of all 19 GABA-A channel subunits in samples of human glioma (n = 29) and peri-tumoral tissue (n = 5). All subunits except the ρ1 and ρ3 subunit were consistently detected. Lowest mRNA levels were found in glioblastoma compared to gliomas of lower malignancy, except for the θ subunit. The expression and cellular distribution of the α1, γ1, ρ2 and θ subunit proteins was investigated by immunohistochemistry on tissue microarrays containing 87 gliomas grade II. We found a strong co-expression of ρ2 and θ subunits in both astrocytomas (r = 0.86, p<0.0001) and oligodendroglial tumors (r = 0.66, p<0.0001). Kaplan-Meier analysis and Cox proportional hazards modeling to estimate the impact of GABA-A channel subunit expression on survival identified the ρ2 subunit (p = 0.043) but not the θ subunit (p = 0.64) as an independent predictor of improved survival in astrocytomas, together with established prognostic factors. Our data give support for the presence of distinct GABA-A channel subtypes in gliomas and provide the first link between specific composition of the A-channel and patient survival

Crystal Structure of the Heteromolecular Chaperone, AscE-AscG, from the Type III Secretion System in Aeromonas hydrophila

10.1371/journal.pone.0019208PLoS ONE64

GABA Maintains the Proliferation of Progenitors in the Developing Chick Ciliary Marginal Zone and Non-Pigmented Ciliary Epithelium

GABA is more than the main inhibitory neurotransmitter found in the adult CNS. Several studies have shown that GABA regulates the proliferation of progenitor and stem cells. This work examined the effects of the GABAA receptor system on the proliferation of retinal progenitors and non-pigmented ciliary epithelial (NPE) cells. qRT-PCR and whole-cell patch-clamp electrophysiology were used to characterize the GABAA receptor system. To quantify the effects on proliferation by GABAA receptor agonists and antagonists, incorporation of thymidine analogues was used. The results showed that the NPE cells express functional extrasynaptic GABAA receptors with tonic properties and that low concentration of GABA is required for a baseline level of proliferation. Antagonists of the GABAA receptors decreased the proliferation of dissociated E12 NPE cells. Bicuculline also had effects on progenitor cell proliferation in intact E8 and E12 developing retina. The NPE cells had low levels of the Cl–transporter KCC2 compared to the mature retina, suggesting a depolarising role for the GABAA receptors. Treatment with KCl, which is known to depolarise membranes, prevented some of the decreased proliferation caused by inhibition of the GABAA receptors. This supported the depolarising role for the GABAA receptors. Inhibition of L-type voltage-gated Ca2+ channels (VGCCs) reduced the proliferation in the same way as inhibition of the GABAA receptors. Inhibition of the channels increased the expression of the cyclin-dependent kinase inhibitor p27KIP1, along with the reduced proliferation. These results are consistent with that when the membrane potential indirectly regulates cell proliferation with hyperpolarisation of the membrane potential resulting in decreased cell division. The increased expression of p27KIP1 after inhibition of either the GABAA receptors or the L-type VGCCs suggests a link between the GABAA receptors, membrane potential, and intracellular Ca2+ in regulating the cell cycle

Evolution of an endofungal Lifestyle: Deductions from the Burkholderia rhizoxinica Genome

<p>Abstract</p> <p>Background</p> <p><it>Burkholderia rhizoxinica </it>is an intracellular symbiont of the phytopathogenic zygomycete <it>Rhizopus microsporus</it>, the causative agent of rice seedling blight. The endosymbiont produces the antimitotic macrolide rhizoxin for its host. It is vertically transmitted within vegetative spores and is essential for spore formation of the fungus. To shed light on the evolution and genetic potential of this model organism, we analysed the whole genome of <it>B. rhizoxinica </it>HKI 0454 - a type strain of endofungal <it>Burkholderia </it>species.</p> <p>Results</p> <p>The genome consists of a structurally conserved chromosome and two plasmids. Compared to free-living <it>Burkholderia </it>species, the genome is smaller in size and harbors less transcriptional regulator genes. Instead, we observed accumulation of transposons over the genome. Prediction of primary metabolic pathways and transporters suggests that endosymbionts consume host metabolites like citrate, but might deliver some amino acids and cofactors to the host. The rhizoxin biosynthesis gene cluster shows evolutionary traces of horizontal gene transfer. Furthermore, we analysed gene clusters coding for nonribosomal peptide synthetases (NRPS). Notably, <it>B. rhizoxinica </it>lacks common genes which are dedicated to quorum sensing systems, but is equipped with a large number of virulence-related factors and putative type III effectors.</p> <p>Conclusions</p> <p><it>B. rhizoxinica </it>is the first endofungal bacterium, whose genome has been sequenced. Here, we present models of evolution, metabolism and tools for host-symbiont interaction of the endofungal bacterium deduced from whole genome analyses. Genome size and structure suggest that <it>B. rhizoxinica </it>is in an early phase of adaptation to the intracellular lifestyle (genome in transition). By analysis of tranporters and metabolic pathways we predict how metabolites might be exchanged between the symbiont and its host. Gene clusters for biosynthesis of secondary metabolites represent novel targets for genomic mining of cryptic natural products. <it>In silico </it>analyses of virulence-associated genes, secreted proteins and effectors might inspire future studies on molecular mechanisms underlying bacterial-fungal interaction.</p

Reduced Secretion of YopJ by Yersinia Limits In Vivo Cell Death but Enhances Bacterial Virulence

Numerous microbial pathogens modulate or interfere with cell death pathways in cultured cells. However, the precise role of host cell death during in vivo infection remains poorly understood. Macrophages infected by pathogenic species of Yersinia typically undergo an apoptotic cell death. This is due to the activity of a Type III secreted effector protein, designated YopJ in Y. pseudotuberculosis and Y. pestis, and YopP in the closely related Y. enterocolitica. It has recently been reported that Y. enterocolitica YopP shows intrinsically greater capacity for being secreted than Y. pestis YopJ, and that this correlates with enhanced cytotoxicity observed for high virulence serotypes of Y. enterocolitica. The enzymatic activity and secretory capacity of YopP from different Y. enterocolitica serotypes have been shown to be variable. However, the underlying basis for differential secretion of YopJ/YopP, and whether reduced secretion of YopJ by Y. pestis plays a role in pathogenesis during in vivo infection, is not currently known. It has also been reported that similar to macrophages, Y. enterocolitica infection of dendritic cells leads to YopP-dependent cell death. We demonstrate here that in contrast to Y. enterocolitica, Y. pseudotuberculosis infection of bone marrow–derived dendritic cells does not lead to increased cell death. However, death of Y. pseudotuberculosis–infected dendritic cells is enhanced by ectopic expression of YopP in place of YopJ. We further show that polymorphisms at the N-terminus of the YopP/YopJ proteins are responsible for their differential secretion, translocation, and consequent cytotoxicity. Mutation of two amino acids in YopJ markedly enhanced both translocation and cytotoxicity. Surprisingly, expression of YopP or a hypersecreted mutant of YopJ in Y. pseudotuberculosis resulted in its attenuation in oral mouse infection. Complete absence of YopJ also resulted in attenuation of virulence, in accordance with previous observations. These findings suggest that control of cytotoxicity is an important virulence property for Y. pseudotuberculosis, and that intermediate levels of YopJ-mediated cytotoxicity are necessary for maximal systemic virulence of this bacterial pathogen