A Possible Role for Short Introns in the Acquisition of Stroma-Targeting Peptides in the Flagellate Euglena gracilis

The chloroplasts of Euglena gracilis bounded by three membranes arose via secondary endosymbiosis of a green alga in a heterotrophic euglenozoan host. Many genes were transferred from symbiont to the host nucleus. A subset of Euglena nuclear genes of predominately symbiont, but also host, or other origin have obtained complex presequences required for chloroplast targeting. This study has revealed the presence of short introns (41–93 bp) either in the second half of presequence-encoding regions or shortly downstream of them in nine nucleus-encoded E. gracilis genes for chloroplast proteins (Eno29, GapA, PetA, PetF, PetJ, PsaF, PsbM, PsbO, and PsbW). In addition, the E. gracilis Pbgd gene contains two introns in the second half of presequence-encoding region and one at the border of presequence-mature peptide-encoding region. Ten of 12 introns present within presequence-encoding regions or shortly downstream of them identified in this study have typical eukaryotic GT/AG borders, are T-rich, 45–50 bp long, and pairwise sequence identities range from 27 to 61%. Thus single recombination events might have been mediated via these cis-spliced introns. A double crossing over between these cis-spliced introns and trans-spliced introns present in 5′-UTRs of Euglena nuclear genes is also likely to have occurred. Thus introns and exon-shuffling could have had an important role in the acquisition of chloroplast targeting signals in E. gracilis. The results are consistent with a late origin of photosynthetic euglenids.     

Characterization of the reduction steps of Fe(III) porphyrins

Polarographic studies have shown that Fe(III) porphyrins undergo successively three one-electron reduction steps in dimethylformamide. The first involves the Fe(III)/Fe(II) redox couple. The second step proceeds to a second reduction of the metal ion and is attributed to the Fe(II)/Fe(I)_couple. This new reduction state of iron porphyrins has been characterized by ESR spectra and by absorption spectra in various solvents. This compound is not axially liganded by strong nucleophilic bases but is sensitive to solvation, the lone electron being localised in the d_z2 orbital. The third reduction step is assumed to involve a reduction of the porphyrin π-electron system.All these results have been confirmed by chemical reductions in tetrahydrofuran.     

Targeted In Vivo Inhibition of Specific Protein–Protein Interactions Using Recombinant Antibodies

With the growing availability of genomic sequence information, there is an increasing need for gene function analysis. Antibody-mediated “silencing” represents an intriguing alternative for the precise inhibition of a particular function of biomolecules. Here, we describe a method for selecting recombinant antibodies with a specific purpose in mind, which is to inhibit intrinsic protein–protein interactions in the cytosol of plant cells. Experimental procedures were designed for conveniently evaluating desired properties of recombinant antibodies in consecutive steps. Our selection method was successfully used to develop a recombinant antibody inhibiting the interaction of ARABIDOPSIS HISTIDINE PHOSPHOTRANSFER PROTEIN 3 with such of its upstream interaction partners as the receiver domain of CYTOKININ INDEPENDENT HISTIDINE KINASE 1. The specific down-regulation of the cytokinin signaling pathway in vivo demonstrates the validity of our approach. This selection method can serve as a prototype for developing unique recombinant antibodies able to interfere with virtually any biomolecule in the living cell.     

RAP: a new computer program for de novo identification of repeated sequences in whole genomes

MOTIVATION: DNA repeats are a common feature of most genomic sequences. Their de novo identification is still difficult despite being a crucial step in genomic analysis and oligonucleotides design. Several efficient algorithms based on word counting are available, but too short words decrease specificity while long words decrease sensitivity, particularly in degenerated repeats. RESULTS: The Repeat Analysis Program (RAP) is based on a new word-counting algorithm optimized for high resolution repeat identification using gapped words. Many different overlapping gapped words can be counted at the same genomic position, thus producing a better signal than the single ungapped word. This results in better specificity both in terms of low-frequency detection, being able to identify sequences repeated only once, and highly divergent detection, producing a generally high score in most intron sequences. AVAILABILITY: The program is freely available for non-profit organizations, upon request to the authors. CONTACT: giorgio.valle@unipd.it SUPPLEMENTARY INFORMATION: The program has been tested on the Caenorhabditis elegans genome using word lengths of 12, 14 and 16 bases. The full analysis has been implemented in the UCSC Genome Browser and is accessible at http://genome.cribi.unipd.it.     

Numerical aspects of optimal control of penicillin production

Since their discovery, fermentation processes have gone along not only with the industrial beverages production and breweries, but since the times of Alexander Fleming, they have become a crucial part of the health care due to antibiotics production. However, complicated dynamics and strong nonlinearities cause that the production with the use of linear control methods achieves only suboptimal yields. From the variety of nonlinear approaches, gradient method has proved the ability to handle these issues—nevertheless, its potential in the field of fermentation processes has not been revealed completely. This paper describes constant vaporization control strategy based on a double-input optimization approach with a successful reduction to a single-input optimization task. To accomplish this, model structure used in the previous work is modified so that it corresponds with the new optimization strategy. Furthermore, choice of search step is explored and various alternatives are evaluated and compared.     

Human tear fluid lipidome: from composition to function

We have explored human aqueous tear fluid lipidome with an emphasis to identify the major lipids. We also address the physiological significance of the lipidome. The tears were analysed using thin layer chromatographic, enzymatic and mass spectrometric techniques. To emphasize the physiological aspect of the lipidome, we modelled the spreading of the non-polar tear fluid lipids at air-water interface in macroscopic scale with olive oil and egg yolk phosphatidylcholine. Based on enzymatic analysis the respective concentrations of choline-containing lipids, triglycerides, and cholesteryl esters were 48±14, 10±0, and 21±18 μM. Ultra performance liquid chromatography quadrupole time of flight mass spectrometry analysis showed that phosphatidylcholine and phosphatidylethanolamine were the two most common polar lipids comprising 88±6% of all identified lipids. Triglycerides were the only non-polar lipids detected in mass spectrometric analysis i.e. no cholesteryl or wax esters were identified. The spreading experiments show that the presence of polar lipids is an absolute necessity for a proper spreading of non-polar tear fluid lipids. We provide evidence that polar lipids are the most common lipid species. Furthermore, we provide a physiological rationale for the observed lipid composition. The results open insights into the functional role of lipids in the tear fluid and also aids in providing new means to understand and treat diseases of the ocular surface.     

Modeling and empirical validation of long‐term carbon sequestration in forests (France, 1850–2015)

The development of appropriate tools to quantify long‐term carbon (C) budgets following forest transitions, that is, shifts from deforestation to afforestation, and to identify their drivers are key issues for forging sustainable land‐based climate‐change mitigation strategies. Here, we develop a new modeling approach, CRAFT (CaRbon Accumulation in ForesTs) based on widely available input data to study the C dynamics in French forests at the regional scale from 1850 to 2015. The model is composed of two interconnected modules which integrate biomass stocks and flows (Module 1) with litter and soil organic C (Module 2) and build upon previously established coupled climate‐vegetation models. Our model allows to develop a comprehensive understanding of forest C dynamics by systematically depicting the integrated impact of environmental changes and land use. Model outputs were compared to empirical data of C stocks in forest biomass and soils, available for recent decades from inventories, and to a long‐term simulation using a bookkeeping model. The CRAFT model reliably simulates the C dynamics during France's forest transition and reproduces C‐fluxes and stocks reported in the forest and soil inventories, in contrast to a widely used bookkeeping model which strictly only depicts C‐fluxes due to wood extraction. Model results show that like in several other industrialized countries, a sharp increase in forest biomass and SOC stocks resulted from forest area expansion and, especially after 1960, from tree growth resulting in vegetation thickening (on average 7.8 Mt C/year over the whole period). The difference between the bookkeeping model, 0.3 Mt C/year in 1850 and 21 Mt C/year in 2015, can be attributed to environmental and land management changes. The CRAFT model opens new grounds for better quantifying long‐term forest C dynamics and investigating the relative effects of land use, land management, and environmental change.     

On the origin of cancer: can we ignore coherence?

A growing number of inconsistencies have accumulated within the genetically deterministic paradigm of the origin of cancer. Among them the most important are the nonspecific nature of cancer mutations and the non-cell-autonomous factors of cancer initiation and progression. Epigenetic aspects of cancer and cancer systems biology represent novel approaches to cancer aetiology and converge in the notion that cancer is characterized by a nonspecific progressive destabilization of multiple molecular pathways. The coherent behaviour of certain cellular subsystems has been theoretically predicted for a long time to have a general role in coordinating biological processes. However, it has only recently gained major scientific interest when it was measured on photosynthetic complexes at physiological temperatures and confirmed to have a direct effect over the dynamics of the energy transfer. Several theoretical and experimental considerations suggest that cancer might be associated with the absence or impairment of the proper coherent dynamics in certain biological structures, most notably in the microtubules. We review those models and suggest that impaired coherence might largely contribute to the progressive destabilization of the molecular and gene regulatory networks, thus connecting different non-genetic aspects of cancer.