Phosphate availability regulates root system architecture in Arabidopsis

Plant root systems are highly plastic in their development and can adapt their architecture in response to the prevailing environmental conditions. One important parameter is the availability of phosphate, which is highly immobile in soil such that the arrangement of roots within the soil will profoundly affect the ability of the plant to acquire this essential nutrient. Consistent with this, the availability of phosphate was found to have a marked effect on the root system architecture of Arabidopsis. Low phosphate availability favored lateral root growth over primary root growth, through increased lateral root density and length, and reduced primary root growth mediated by reduced cell elongation. The ability of the root system to respond to phosphate availability was found to be independent of sucrose supply and auxin signaling. In contrast, shoot phosphate status was found to influence the root system architecture response to phosphate availability

Bioinformatics prediction of overlapping frameshifted translation products in mammalian transcripts

<p>Abstract</p> <p>Background</p> <p>Exceptionally, a single nucleotide sequence can be translated <it>in vivo </it>in two different frames to yield distinct proteins. In the case of the G-protein alpha subunit XL-alpha-s transcript, a frameshifted open reading frame (ORF) in exon 1 is translated to yield a structurally distinct protein called Alex, which plays a role in platelet aggregation and neurological processes. We carried out a novel bioinformatics screen for other possible dual-frame translated sequences, based on comparative genomics.</p> <p>Results</p> <p>Our method searched human, mouse and rat transcripts in frames +1 and -1 for ORFs which are unusually well conserved at the amino acid level. We name these conserved frameshifted overlapping ORFs 'matreshkas' to reflect their nested character. Select findings of our analysis revealed that the G-protein coupled receptor GPR27 is entirely contained within a frame -1 matreshka, thrombopoietin contains a matreshka which spans ~70% of its length, platelet glycoprotein IIIa (ITGB3) contains a matreshka with the predicted characteristics of a secreted peptide hormone, while the potassium channel KCNK12 contains a matreshka spanning >400 amino acids.</p> <p>Conclusion</p> <p>Although the <it>in vivo </it>existence of translated matreshkas has not been experimentally verified, this genome-wide analysis provides strong evidence that substantial overlapping coding sequences exist in a number of human and rodent transcripts.</p

HAltORF: a database of predicted out-of-frame alternative open reading frames in human

Human alternative open reading frames (HAltORF) is a publicly available and searchable online database referencing putative products of out-of-frame alternative translation initiation (ATI) in human mRNAs. Out-of-frame ATI is a process by which a single mRNA encodes independent proteins, when distinct initiation codons located in different reading frames are recognized by a ribosome to initiate translation. This mechanism is largely used in viruses to increase the coding potential of small viral genomes. There is increasing evidence that out-of-frame ATI is also used in eukaryotes, including human, and may contribute to the diversity of the human proteome. HAltORF is the first web-based searchable database that allows thorough investigation in the human transcriptome of out-of-frame alternative open reading frames with a start codon located in a strong Kozak context, and are thus the more likely to be expressed. It is also the first large scale study on the human transcriptome to successfully predict the expression of out-of-frame ATI protein products that were previously discovered experimentally. HAltORF will be a useful tool for the identification of human genes with multiple coding sequences, and will help to better define and understand the complexity of the human proteome

Phosphate control of root system architecture in Arabidopsis

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Use of nanogold- and fluorescent-labeled antibody Fv fragments in immunocytochemistry

Recombinant antibody fragments are emerging as a versatile tool in both basic research and medical therapy. We describe the procedures for direct labeling of engineered antibody fragments (Fv) with fluorescein or nanogold and their use in fluorescence and immunoelectron microscopy, respectively. The Fv fragments were produced in Escherichia coli, purified by one-step Strep tag affinity chromatography, chemically labeled with the marker, and employed in microscopy to localize epitopes on the membrane protein bacteriorhodopsin in purple membranes of Halobacterium halobium and the cytochrome c oxidase of Paracoccus denitrificans. In both cases, methods involving directly labeled antibody fragments show results identical to those in which antibodies or Fv fragments are detected by a secondarily labeled conjugate. The multifunctional design of the recombinant Fv fragments, however, offers more all-around applications in immunocytochemistry. The directly labeled Fv fragments, half the size of an Fab fragment, are at the molecular level the smallest antibody fragments yet described for visualization of biomolecules in microscopy

Alternative Complement Pathway Inhibition Abrogates Pneumococcal Opsonophagocytosis in Vaccine-Naïve, but Not in Vaccinated Individuals

To assess the relative contribution of opsonisation by antibodies, classical and alternative complement pathways to pneumococcal phagocytosis, we analyzed killing of pneumococci by human blood leukocytes collected from vaccine-naïve and PCV13-vaccinated subjects. With serotype 4 pneumococci as model, two different physiologic opsonophagocytosis assays based on either hirudin-anticoagulated whole blood or on washed cells from EDTA-anticoagulated blood reconstituted with active serum, were compared. Pneumococcal killing was measured in the presence of inhibitors targeting the complement components C3, C5, MASP-2, factor B or factor D. The two assay formats yielded highly consistent and comparable results. They highlighted the importance of alternative complement pathway activation for efficient opsonophagocytic killing in blood of vaccine-naïve subjects. In contrast, alternative complement pathway inhibition did not affect pneumococcal killing in PCV13-vaccinated individuals. Independent of amplification by the alternative pathway, even low capsule-specific antibody concentrations were sufficient to efficiently trigger classical pathway mediated opsonophagocytosis. In heat-inactivated or C3-inhibited serum, high concentrations of capsule-specific antibodies were required to trigger complement-independent opsonophagocytosis. Our findings suggest that treatment with alternative complement pathway inhibitors will increase susceptibility for invasive pneumococcal infection in non-immune subjects, but it will not impede pneumococcal clearance in vaccinated individuals

Bioinformatics prediction of overlapping frameshifted translation products in mammalian transcripts-0

Ike proteins which start with methionine, a first run of the analysis pipeline generated redundant (overlapping) methionine-start ORFs. A second, more comprehensive run generated all ORFs regardless of the first amino acid.<p><b>Copyright information:</b></p><p>Taken from "Bioinformatics prediction of overlapping frameshifted translation products in mammalian transcripts"</p><p>http://www.biomedcentral.com/1471-2164/9/122</p><p>BMC Genomics 2008;9():122-122.</p><p>Published online 6 Mar 2008</p><p>PMCID:PMC2329644.</p><p></p

Bioinformatics prediction of overlapping frameshifted translation products in mammalian transcripts-3

Ercase, Kozak motifs in bold. The bold white guanine base (grey background) can be replaced by adenine to truncate the matreshka through a SNP (NCBI ID rs1126665), and replaces a glycine with a glutamate within the Erythropoietin-Thrombopoietin motif (Pfam ID PF00758.7). Alternating italics/non-italics represent alternate THPO coding exons, and spaces separate codons for the parent RefSeq. B) matreshka protein sequence. C) Illustration of the translation of the THPO matreshka in the nirs splice variant. THPO splice variant 1 is shown above the nirs splice variant. Hatching indicates the location of the matreshka. Nirs splicing utilises an alternative acceptor which results in the last exon being translated in the same frame as the matreshka over 564 nucleotides (indicated by grey shading).<p><b>Copyright information:</b></p><p>Taken from "Bioinformatics prediction of overlapping frameshifted translation products in mammalian transcripts"</p><p>http://www.biomedcentral.com/1471-2164/9/122</p><p>BMC Genomics 2008;9():122-122.</p><p>Published online 6 Mar 2008</p><p>PMCID:PMC2329644.</p><p></p