Regulation of Surface Proteins Assembly on the Wall of Gram-Positive Bacteria

Cell wall peptidoglycan-anchored surface proteins are essential virulence factors in many Gram-positive bacteria. Attachment of these proteins to the peptidoglycan is achieved through a transpeptidation reaction, whereby sortase cleaves a conserved Cterminal LPXTG motif and attaches the protein to the peptidoglycan precursor lipid II. This thesis deals with the spatial regulation of the sorting reaction, and the role sortase localization plays in the correct distribution of surface proteins. We have introduced a new immunofluorescence procedure to study the distribution of sub-surface antigens in Streptococcus pyogenes, which utilizes the phage lysin PlyC to permeabilize the cell wall of to antibodies. We found that sortase localizes within distinct membranal foci, the majority of which are associated with the division septum and colocalize with areas of active M-protein anchoring. Protein anchoring takes place at two distinct cellular locations, the division septum and the poles. Anchoring of M-protein at the septum is a rapid process that occurs in concert with septal peptidoglycan synthesis. The localized secretion of M-protein, as well as the prevalence of sortase, lipid II, and PBPs at this location, promote efficient protein anchoring. Sortase localization and M-protein anchoring occurs simultaneously at the septum and the mature equatorial rings, pointing to the possibility that cell division begins in the daughter cells before the conclusion of the previous division cycle. Anchoring of SfbI at the poles is a much slower and more diffuse process, and therefore requires less sortase. The protein sorting reaction is therefore a dynamic, and a highly regulated process. In the absence of sortase A, surface proteins accumulate at the membrane-wall interface. We found that stalled surface proteins can be released from the secretion channel by an LPXTG-specific enzyme other than sortase A, whose identity is still unknown. Nonetheless, accumulation of missorted surface proteins has deleterious effects on the cell, resulting in selective pressure to repress surface protein expression. Inhibition of sortase may therefore not only prevent surface protein anchoring but also exert direct pressure on the cell’s secretion and folding pathways. A better understanding of the mechanisms controlling the biogenesis of surface molecules, aided by the ability to study sub-surface antigens through immunofluorescence, may yield promising new candidates for the development of new anti-infecting agents

Regulation of Zn and Fe transporters by the GPC1 gene during early wheat monocarpic senescence

BACKGROUND: During wheat senescence, leaf components are degraded in a coordinated manner, releasing amino acids and micronutrients which are subsequently transported to the developing grain. We have previously shown that the simultaneous downregulation of Grain Protein Content (GPC) transcription factors, GPC1 and GPC2, greatly delays senescence and disrupts nutrient remobilization, and therefore provide a valuable entry point to identify genes involved in micronutrient transport to the wheat grain. RESULTS: We generated loss-of-function mutations for GPC1 and GPC2 in tetraploid wheat and showed in field trials that gpc1 mutants exhibit significant delays in senescence and reductions in grain Zn and Fe content, but that mutations in GPC2 had no significant effect on these traits. An RNA-seq study of these mutants at different time points showed a larger proportion of senescence-regulated genes among the GPC1 (64%) than among the GPC2 (37%) regulated genes. Combined, the two GPC genes regulate a subset (21.2%) of the senescence-regulated genes, 76.1% of which are upregulated at 12 days after anthesis, before the appearance of any visible signs of senescence. Taken together, these results demonstrate that GPC1 is a key regulator of nutrient remobilization which acts predominantly during the early stages of senescence. Genes upregulated at this stage include transporters from the ZIP and YSL gene families, which facilitate Zn and Fe export from the cytoplasm to the phloem, and genes involved in the biosynthesis of chelators that facilitate the phloem-based transport of these nutrients to the grains. CONCLUSIONS: This study provides an overview of the transport mechanisms activated in the wheat flag leaf during monocarpic senescence. It also identifies promising targets to improve nutrient remobilization to the wheat grain, which can help mitigate Zn and Fe deficiencies that afflict many regions of the developing world.Fil: Pearce, Stephen. University of California; Estados UnidosFil: Tabbita, Facundo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Recursos Naturales. Instituto de Recursos Biológicos; ArgentinaFil: Cantu, Dario. University of California; Estados UnidosFil: Buffalo, Vince. University of California; Estados UnidosFil: Avni, Raz. Tel Aviv University; IsraelFil: Vazquez Gross, Hans. University of California; Estados UnidosFil: Zhao, Rongrong. China Agricultural University; ChinaFil: Conley, Christopher J.. University of California; Estados UnidosFil: Distelfeld, Assaf. Faculty Of Life Sciences, Department Of Molecular Biolo;Fil: Dubcovsky, Jorge. University of California; Estados Unidos. Howard Hughes Medical Institute ; Estados Unidos. Gordon & Betty Moore Foundation Investigator; Estados Unido

Is IL-1β Further Evidence for the Role of Propionibacterium acnes in Degenerative Disc Disease? Lessons From the Study of the Inflammatory Skin Condition Acne Vulgaris

The pathogenesis of degenerative disc disease is a complex and multifactorial process in which genetics, mechanical trauma, altered loading and nutrition present significant etiological factors. Infection of the intervertebral disc with the anaerobic bacterium Propionibacterium acnes is now also emerging as a potentially new etiological factor. This human commensal bacterium is well known for its long association with the inflammatory skin condition acne vulgaris. A key component of inflammatory responses to P. acnes in acne appears to be interleukin (IL)-1β. Similarly, in degenerative disc disease (DDD) there is compelling evidence for the fundamental roles of IL-1β in its pathology. We therefore propose that P. acnes involvement in DDD is biologically very plausible, and that IL-1β is the key inflammatory mechanism driving the host response to P. acnes infection. Since there is a solid theoretical basis for this phenomenon, we further propose that the relationship between P. acnes infection and DDD is causal

Importance of Propionibacterium acnes hemolytic activity in human intervertebral discs:A microbiological study

Most patients with chronic lower back pain (CLBP) exhibit degenerative disc disease. Disc specimens obtained during initial therapeutic discectomies are often infected/colonized with Propionibacterium acnes, a Gram-positive commensal of the human skin. Although pain associated with infection is typically ascribed to the body's inflammatory response, the Gram-positive bacterium Staphylococcus aureus was recently observed to directly activate nociceptors by secreting pore-forming α-hemolysins that disrupt neuronal cell membranes. The hemolytic activity of P. acnes in cultured disc specimens obtained during routine therapeutic discectomies was assessed through incubation on sheep-blood agar. The β-hemolysis pattern displayed by P. acnes on sheep-blood agar was variable and phylogroup-dependent. Their molecular phylogroups were correlated with their hemolytic patterns. Our findings raise the possibility that pore-forming proteins contribute to the pathogenesis and/or symptomology of chronic P. acnes disc infections and CLBP, at least in a subset of cases

Cation Diffusion Facilitators Transport Initiation and Regulation Is Mediated by Cation Induced Conformational Changes of the Cytoplasmic Domain

Cation diffusion facilitators (CDF) are part of a highly conserved protein family that maintains cellular divalent cation homeostasis in all domains of life. CDF's were shown to be involved in several human diseases, such as Type-II diabetes and neurodegenerative diseases. In this work, we employed a multi-disciplinary approach to study the activation mechanism of the CDF protein family. For this we used MamM, one of the main ion transporters of magnetosomes - bacterial organelles that enable magnetotactic bacteria to orientate along geomagnetic fields. Our results reveal that the cytosolic domain of MamM forms a stable dimer that undergoes distinct conformational changes upon divalent cation binding. MamM conformational change is associated with three metal binding sites that were identified and characterized. Altogether, our results provide a novel auto-regulation mode of action model in which the cytosolic domain's conformational changes upon ligand binding allows the priming of the CDF into its transport mode

Differential Development of Human Brain White Matter Tracts

Neuroscience is increasingly focusing on developmental factors related to human structural and functional connectivity. Unfortunately, to date, diffusion-based imaging approaches have only contributed modestly to these broad objectives, despite the promise of diffusion-based tractography. Here, we report a novel data-driven approach to detect similarities and differences among white matter tracts with respect to their developmental trajectories, using 64-direction diffusion tensor imaging. Specifically, using a cross-sectional sample comprising 144 healthy individuals (7 to 48 years old), we applied k-means cluster analysis to separate white matter voxels based on their age-related trajectories of fractional anisotropy. Optimal solutions included 5-, 9- and 14-clusters. Our results recapitulate well-established tracts (e.g., internal and external capsule, optic radiations, corpus callosum, cingulum bundle, cerebral peduncles) and subdivisions within tracts (e.g., corpus callosum, internal capsule). For all but one tract identified, age-related trajectories were curvilinear (i.e., inverted ‘U-shape’), with age-related increases during childhood and adolescence followed by decreases in middle adulthood. Identification of peaks in the trajectories suggests that age-related losses in fractional anisotropy occur as early as 23 years of age, with mean onset at 30 years of age. Our findings demonstrate that data-driven analytic techniques may be fruitfully applied to extant diffusion tensor imaging datasets in normative and neuropsychiatric samples

Making cheaper labor: Domestic outsourcing and development in the Galilee

Middle Eastern Studie