Impaired Mitochondrial Microbicidal Responses in Chronic Obstructive Pulmonary Disease Macrophages

RATIONALE: Chronic obstructive pulmonary disease (COPD) is characterized by impaired clearance of pulmonary bacteria. OBJECTIVES: The effect of COPD on alveolar macrophage (AM) microbicidal responses was investigated. METHODS: Alveolar macrophages (AMs) were obtained from bronchoalveolar lavage from healthy donors or COPD patients and challenged with opsonized serotype 14 Streptococcus pneumoniae. Cells were assessed for apoptosis, bactericidal activity and mitochondrial reactive oxygen species (mROS) production. A transgenic mouse line, in which the CD68 promoter ensures macrophage specific expression of human Mcl-1 (CD68.hMcl-1), was used to model the molecular aspects of COPD. MEASUREMENTS AND MAIN RESULTS: COPD AM had elevated levels of Mcl-1, an anti-apoptotic Bcl-2 family member, with selective reduction of delayed intracellular bacterial killing. CD68.hMcl-1 AM phenocopied the microbicidal defect since transgenic mice demonstrated impaired clearance of pulmonary bacteria and increased neutrophilic inflammation. Murine bone marrow-derived macrophages (BMDM) and human monocyte-derived macrophages (MDM) generated mitochondrial reactive oxygen species (mROS) in response to pneumococci, which co-localized with bacteria and phagolysosomes to enhance bacterial killing. The Mcl-1 transgene increased oxygen consumption rates and mROS expression in mock-infected BMDM but reduced caspase-dependent mROS production after pneumococcal challenge. COPD AM also increased basal mROS expression, but failed to increase production after pneumococcal challenge, in keeping with reduced intracellular bacterial killing. The defect in COPD AM intracellular killing was associated with a reduced ratio of mROS /superoxide dismutase 2. CONCLUSIONS: Upregulation of Mcl-1 and chronic adaption to oxidative stress alters mitochondrial metabolism and microbicidal function, reducing the delayed phase of intracellular bacterial clearance in COPD

Alteration of the embryo transcriptome of hexaploid winter wheat (Triticum aestivum cv. Mercia) during maturation and germination

Grain dormancy and germination are areas of biology that are of considerable interest to the cereal community. We have used a 9,155-feature wheat unigene cDNA microarray resource to investigate changes in the wheat embryo transcriptome during late grain development and maturation and during the first 48 h of postimbibition germination. In the embryo 392 mRNAs accumulated by twofold or greater over the time course from 21 days postanthesis (dpa) to 40 dpa and on through 1 and 2 days postgermination. These included mRNAs encoding proteins involved in amino acid biosynthesis and metabolism, cell division and subsequent cell development, signal transduction, lipid metabolism, energy production, protein turnover, respiration, initiation of transcription, initiation of translation and ribosomal composition. A number of mRNAs encoding proteins of unknown function also accumulated over the time course. Conversely 163 sequences showed decreases of twofold or greater over the time course. A small number of mRNAs also showed rapid accumulation specifically during the first 48 h of germination. We also examined alterations in the accumulation of transcripts encoding proteins involved in abscisic acid signalling. Thus, we describe changes in the level of transcripts encoding wheat Viviparous 1 (Vp1) and other interacting proteins. Interestingly, the transcript encoding wheat Viviparous-interacting protein 1 showed a pattern of accumulation that correlates inversely with germination. Our data suggests that the majority of the transcripts required for germination accumulate in the embryo prior to germination and we discuss the implications of these findings with regard to manipulation of germination in wheat

Helicity within the vortex filament model

Kinetic helicity is one of the invariants of the Euler equations that is associated with the topology of vortex lines within the fluid. In superfluids, the vorticity is concentrated along vortex filaments. In this setting, helicity would be expected to acquire its simplest form. However, the lack of a core structure for vortex filaments appears to result in a helicity that does not retain its key attribute as a quadratic invariant. By defining a spanwise vector to the vortex through the use of a Seifert framing, we are able to introduce twist and henceforth recover the key properties of helicity. We present several examples for calculating internal twist to illustrate why the centreline helicity alone will lead to ambiguous results if a twist contribution is not introduced. Our choice of the spanwise vector can be expressed in terms of the tangential component of velocity along the filament. Since the tangential velocity does not alter the configuration of the vortex at later times, we are able to recover a similar equation for the internal twist angle to that of classical vortex tubes. Our results allow us to explain how a quasi-classical limit of helicity emerges from helicity considerations for individual superfluid vortex filaments

Three endo-β-mannanase genes expressed in the micropylar endosperm and in the radicle influence germination of Arabidopsis thaliana seeds

Mannans are hemicellulosic polysaccharides in the plant primary cell wall (CW). Mature seeds, specially their endosperm cells, have CWs rich in mannan-based polymers that confer a strong mechanical resistance for the radicle protrusion upon germination. The rupture of the seed coat and endosperm are two sequential events during the germination of Arabidopsis thaliana. Endo-β-mannanases (MAN; EC. 3.2.1.78) are hydrolytic enzymes that catalyze cleavage of β1 → 4 bonds in the mannan-polymer. In the genome of Arabidopsis, the endo-β-mannanase (MAN) family is represented by eight members. The expression of these eight MAN genes has been systematically explored in different organs of this plant and only four of them (AtMAN7, AtMAN6, AtMAN2 and AtMAN5) are expressed in the germinating seeds. Moreover, in situ hybridization analysis shows that their transcript accumulation is restricted to the micropylar endosperm and to the radicle and this expression disappears soon after radicle emergence. T-DNA insertion mutants in these genes (K.O. MAN7, K.O. MAN6, K.O. MAN5), except that corresponding to AtMAN2 (K.O. MAN2), germinate later than the wild type (Wt). K.O. MAN6 is the most affected in the germination time course with a t 50 almost double than that of the Wt. These data suggest that AtMAN7, AtMAN5 and specially AtMAN6 are important for the germination of A. thaliana seeds by facilitating the hydrolysis of the mannan-rich endosperm cell walls

Crystal Structure of Reovirus Attachment Protein σ1 in Complex with Sialylated Oligosaccharides

Many viruses attach to target cells by binding to cell-surface glycans. To gain a better understanding of strategies used by viruses to engage carbohydrate receptors, we determined the crystal structures of reovirus attachment protein σ1 in complex with α-2,3-sialyllactose, α-2,6-sialyllactose, and α-2,8-di-siallylactose. All three oligosaccharides terminate in sialic acid, which serves as a receptor for the reovirus serotype studied here. The overall structure of σ1 resembles an elongated, filamentous trimer. It contains a globular head featuring a compact β-barrel, and a fibrous extension formed by seven repeating units of a triple β-spiral that is interrupted near its midpoint by a short α -helical coiled coil. The carbohydrate-binding site is located between β-spiral repeats two and three, distal from the head. In all three complexes, the terminal sialic acid forms almost all of the contacts with σ1 in an identical manner, while the remaining components of the oligosaccharides make little or no contacts. We used this structural information to guide mutagenesis studies to identify residues in σ1 that functionally engage sialic acid by assessing hemagglutination capacity and growth in murine erythroleukemia cells, which require sialic acid binding for productive infection. Our studies using σ1 mutant viruses reveal that residues 198, 202, 203, 204, and 205 are required for functional binding to sialic acid by reovirus. These findings provide insight into mechanisms of reovirus attachment to cell-surface glycans and contribute to an understanding of carbohydrate binding by viruses. They also establish a filamentous, trimeric carbohydrate-binding module that could potentially be used to endow other trimeric proteins with carbohydrate-binding properties

The Re-Establishment of Desiccation Tolerance in Germinated Arabidopsis thaliana Seeds and Its Associated Transcriptome

The combination of robust physiological models with “omics” studies holds promise for the discovery of genes and pathways linked to how organisms deal with drying. Here we used a transcriptomics approach in combination with an in vivo physiological model of re-establishment of desiccation tolerance (DT) in Arabidopsis thaliana seeds. We show that the incubation of desiccation sensitive (DS) germinated Arabidopsis seeds in a polyethylene glycol (PEG) solution re-induces the mechanisms necessary for expression of DT. Based on a SNP-tile array gene expression profile, our data indicates that the re-establishment of DT, in this system, is related to a programmed reversion from a metabolic active to a quiescent state similar to prior to germination. Our findings show that transcripts of germinated seeds after the PEG-treatment are dominated by those encoding LEA, seed storage and dormancy related proteins. On the other hand, a massive repression of genes belonging to many other classes such as photosynthesis, cell wall modification and energy metabolism occurs in parallel. Furthermore, comparison with a similar system for Medicago truncatula reveals a significant overlap between the two transcriptomes. Such overlap may highlight core mechanisms and key regulators of the trait DT. Taking into account the availability of the many genetic and molecular resources for Arabidopsis, the described system may prove useful for unraveling DT in higher plants

Effects of APETALA2 on embryo, endosperm, and seed coat development determine seed size in Arabidopsis

Arabidopsis APETALA2 (AP2) controls seed mass maternally, with ap2 mutants producing larger seeds than wild type. Here, we show that AP2 influences development of the three major seed compartments: embryo, endosperm, and seed coat. AP2 appears to have a significant effect on endosperm development. ap2 mutant seeds undergo an extended period of rapid endosperm growth early in development relative to wild type. This early expanded growth period in ap2 seeds is associated with delayed endosperm cellularization and overgrowth of the endosperm central vacuole. The subsequent period of moderate endosperm growth is also extended in ap2 seeds largely due to persistent cell divisions at the endosperm periphery. The effect of AP2 on endosperm development is mediated by different mechanisms than parent-of-origin effects on seed size observed in interploidy crosses. Seed coat development is affected; integument cells of ap2 mutants are more elongated than wild type. We conclude that endosperm overgrowth and/or integument cell elongation create a larger postfertilization embryo sac into which the ap2 embryo can grow. Morphological development of the embryo is initially delayed in ap2 compared with wild-type seeds, but ap2 embryos become larger than wild type after the bent-cotyledon stage of development. ap2 embryos are able to fill the enlarged postfertilization embryo sac, because they undergo extended periods of cell proliferation and seed filling. We discuss potential mechanisms by which maternally acting AP2 influences development of the zygotic embryo and endosperm to repress seed size