Acid digestion and symbiont: proton sharing at the origin of mitochondriogenesis?: proton production by a symbiotic bacterium may have been the origin of two hallmark eukaryotic features, acid digestion and mitochondria

The initial relationships between organisms leading to endosymbiosis and the first eukaryote are currently a topic of hot debate. Here, I present a theory that offers a gradual scenario in which the origins of phagocytosis and mitochondria are intertwined in such a way that the evolution of one would not be possible without the other. In this scenario, the premitochondrial bacterial symbiont became initially associated with a protophagocytic host on the basis of cooperation to kill prey with symbiontproduced toxins and reactive oxygen species (ROS). Subsequently, the cooperation was focused on the digestion stage, through the acidification of the protophagocytic cavities via exportation of protons produced by the aerobic respiration of the symbiont. The host gained an improved phagocytic capacity and the symbiont received organic compounds from prey. As the host gradually lost its membrane energetics to develop lysosomal digestion, respiration was centralized in the premitochondrial symbiont for energy production for the consortiu

Occupational shoes and podiatric actuation

En el siguiente estudio, tras hacer una presunción sobre el uso del calzado laboral y la patología derivada del uso y el abuso del mismo, el uso del calzado laboral a tenor de evitar traumatismos directos en el pie y la unidad ungueal y otros riesgos, tanto químicos como biológicos y ambientales, también es un factor desencadenante en la producción de la patología podológica. La falta de información que se lleva a cabo en las empresas, en parte por falta de podólogos experimentados en la materia que instruyan a la población laboral en cada empresa, y en parte por falta de interés por el colectivo podológico en abordar la salud podológica laboral como una más de sus funciones, de las cuales podemos producir un importante aumento cualitativo y cuantitativo en la eficacia y eficiencia de los trabajadores y contribuir a introducir el colectivo podológico como parte de los servicios de prevención en las empresas, donde no se evidencia nuestra presencia, también sería importante reseñar la poca coordinación entre las empresas manufactureras de calzados y el muy beneficioso aporte de la podología a estas industrias. A través del estudio realizado a distintos trabajadores de distintas empresas y sectores, he seleccionado información sobre la presencia de patología podológica por uso de calzado laboral y también información sobre la confortabilidad del mismo para delimitar cuales deberían de ser las funciones de la podología en el ámbito laboral.In the next study, before make a presumption about the use and abuse of work shoe and the podiatric pathology, the use of work shoe to prevent of the direct trauma in the foot and nail and other risks, so much chemical risks and biological and environmental risks, as well is a causing factor in the production of the podiatric pathology. The abscense of information in the business, in part for the abscense of experimented podiatrics in this field that learning to the laboral population in the firm, as well for the staff of interest for the podiatric group in the treatment of the podiatric work health as a of the role as podiatry , derive of this functions to produce a important rise as qualitative as quantitative in the effi cacy and the efficiency of the workers in the firms, where there isn´t our presence, also it is very important to review the little coordination between the firms manufacturer of shoes and the best contribution that the podiatry could to make in these fi rms. As well as trough the study done to many workers of many firms and areas, I´ve choosen information about the presence or abscense of podiatric pathology for use of the occupational footwear and also about the confortability for to define who will must to be the duties of podiatric medical in the work area

The main early and late promoters of Bacillus subtilis phage ø29 form unstable open complexes with sA-RNA polymerase that are stabilized by DNA supercoiling

Most Escherichia coli promoters studied so far form stable open complexes with σ70-RNA polymerase which have relatively long half-lives and, therefore, are resistant to a competitor challenge. A few exceptions are nevertheless known. The analysis of a number of promoters in Bacillus subtilis has suggested that the instability of open complexes formed by the vegetative σA polymerase may be a more general phenomenon than in Escherichia coli. We show that the main early and late promoters from the Bacillus subtilis phage ø29 form unstable open complexes that are stabilized either by the formation of the first phosphodlester bond between the initiating nucleoside triphosphates or by DNA supercoiling. The functional characteristics of these two strong promoters suggest that they are not optimized for a tight and stable RNA polymerase binding. Their high activity is probably the consequence of the efficiency of further steps leading to the formation of an elongation complex.This investigation was aided by research grants from the National Institutes of Health (5ROI GM27242-13), Direccion General de Investigaci6n Cientifica y Tecnica (PB90 0091) and an institutional Grant from Fundacidn Ram6n Areces. F.R. was holder of a post-doctoral fellowship from M.E.C. B.N. and M.M were holders of pre-doctoral fellowships from M.E.C and Gobierno Vasco, respectively.Peer reviewe

ICETh1 and ICETh2, two interdependent mobile genetic elements in Thermus thermophilus transjugation.

Cell to cell DNA transfer between Thermus thermophilus, or transjugation, requires the natural competence apparatus (NCA) of the recipient cell and a DNA donation machinery in the donor. In T. thermophilus HB27, two mobile genetic elements with functional similarities to Integrative and Conjugative Elements (ICEs) coexist, ICETh1 encoding the DNA transfer apparatus and ICETh2, encoding a putative replication module. Here, we demonstrate that excision and integration of both elements depend on a single tyrosine recombinase encoded by ICETh2, and that excision is not required but improves the transfer of these elements to a recipient cell. These findings along with previous results suggest that ICETh1 and ICETh2 depend on each other for spreading among T. thermophilus by transjugation.post-print1,28 M

Nuclear localization signals in phage terminal proteins provide a novel gene delivery tool in mammalian cells

Terminal proteins (TPs) of bacteriophages prime DNA replication and become covalently linked to the genome ends. Unexpectedly, we have found functional eukaryotic nuclear localization signals (NLSs) within the TP sequences of bacteriophages from diverse families and hosts. Given the role of bacteriophages as vehicles for horizontal gene transfer (HGT), we postulated that viral genomes that have covalently linked NLS-containing terminal proteins might behave as vectors for HGT between bacteria and the eukaryotic nucleus. To validate this hypothesis, we profited from the in vitro Φ29 amplification system that allows the amplification of heterologous DNAs producing linear molecules of DNA with TP covalently attached to both 5' ends. Interestingly, these in vitrogenerated TP-DNA molecules showed enhanced gene delivery in mammalian cells, supporting a possible role in HGT by transferring genes between prokaryotes and eukaryotes. Moreover, these TP-DNA molecules are a useful tool to amplify and subsequently deliver genes efficiently into the eukaryotic nucleus. Here, we suggest various possible applications and further developments of the technique with biotechnological and therapeutic purposesThis work was supported by Grant BFU 2011-23645 and Consolider-Ingenio Grant 2010 24717 from the Spanish Ministry of Economy and Competitiveness (MINECO) and by an institutional grant from Fundacion Ramon Areces to the Centro de Biologia Molecular “Severo Ochoa.

A modular vector toolkit with a tailored set of thermosensors to regulate gene expression in thermus thermophilus

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acsomega.9b02107Modular plasmid architectures have shown to be a very useful resource to standardize, build, share, and compare biological parts and functional vectors, and are being applied in an increasing number of microorganisms. Here, we present a modular plasmid toolkit for Thermus thermophilus, a species considered as a workhorse for biotechnology and a model for high-temperature biology. Apart from integrating improved versions of already existing parts, we have characterized specific promoters and developed a thermosensor-based palette that restricts the expression to Thermus and, at the same time, controls protein expression in this organism in a temperature-dependent mannerThis work was supported by the European Union grant H2020-BIOTEC-2014-2015/H2020-LEIT-BIO-2015-1 and grant BIO2016-77031R from the Spanish Ministry of Science, Innovation and Universities (MICIU). An institutional grant to CBMSO from Fundación Ramón Areces and an FPI contract to C.V. from the MCIU are also acknowledge

Integrative and Conjugative Element ICETh1 Functions as a Pangenomic DNA Capture Module in Thermus thermophilus.

Transjugation is an unconventional conjugation mechanism in Thermus thermophilus (Tth) that involves the active participation of both mating partners, encompassing a DNA secretion system (DSS) in the donor and an active natural competence apparatus (NCA) in the recipient cells. DSS is encoded within an integrative and conjugative element (ICETh1) in the strain Tth HB27, whereas the NCA is constitutively expressed in both mates. Previous experiments suggested the presence of multiple origins of transfer along the genome, which could generate genomic mosaicity among the progeny. Here, we designed transjugation experiments between two closely related strains of Tth with highly syntenic genomes, containing enough single nucleotide polymorphisms to allow precise parenthood analysis. Individual clones from the progeny were sequenced, revealing their origin as derivatives of our ICETh1-containing intended “donor” strain (HB27), which had acquired separate fragments from the genome of the ICETh1-free HB8 cells, which are our intended recipient. Due to the bidirectional nature of transjugation, only assays employing competence-defective HB27 derivatives as donors allowed the recovery of HB8-derived progeny. These results show a preference for a retrotransfer mechanism in transjugation in ICETh1-bearing strains, supporting an inter-strain gene-capture function for ICETh1. This function could benefit the donor-capable host by facilitating the acquisition of adaptive traits from external sources, ultimately increasing the open pangenome of Thermus, maximizing the potential repertoire of physiological and phenotypical traits related to adaptation and speciation.post-print11136 K

Deletion of the primase-polymerases encoding gene, located in a mobile element in Thermus thermophilus HB27, leads to loss of function mutation of addAB genes

DNA primase-polymerases (Ppol) have been shown to play active roles in DNA repair and damage tolerance, both in prokaryotes and eukaryotes. The ancestral thermophilic bacterium Thermus thermophilus strain HB27 encodes a Ppol protein among the genes present in mobile element ICETh2, absent in other T. thermophilus strains. Using different strategies we ablated the function of Ppol in HB27 cells, either by knocking out the gene through insertional mutagenesis, markerless deletion or through abolition of its catalytic activity. Whole genome sequencing of this diverse collection of Ppol mutants showed spontaneous loss of function mutation in the helicase-nuclease AddAB in every ppol mutant isolated. Given that AddAB is a major player in recombinational repair in many prokaryotes, with similar activity to the proteobacterial RecBCD complex, we have performed a detailed characterization of the ppol mutants in combination with addAB mutants. The results show that knockout addAB mutants are more sensitive to DNA damage agents than the wild type, and present a dramatic three orders of magnitude increase in natural transformation efficiencies with both plasmid and lineal DNA, whereas ppol mutants show defects in plasmid stability. Interestingly, DNA-integrity comet assays showed that the genome of all the ppol and/or addAB mutants was severely affected by widespread fragmentation, however, this did not translate in neat loss of viability of the strains. All these data support that Ppol appears to keep in balance the activity of AddAB as a part of the DNA housekeeping maintenance in T. thermophilus HB27, thus, playing a key role in its genome stabilityThis work was supported by a grant from the Spanish Ministry of Science and Innovation PID2019-109073RB-I00. CV was a holder of a competitive FPI contract. An institutional grant from Fundación Ramón Areces to the CBMSO was also acknowledge

A thermostable DNA primase-polymerase from a mobilegenetic element involved in defence againstenvironmental DNA.

Primase-polymerases (Ppol) are one of the few enzymes able to start DNA synthesis on ssDNA templates. The role of Thermus thermophilus HB27 Ppol, encoded along a putative helicase (Hel) within a mobile genetic element (ICETh2), has been studied. A mutant lacking Ppol showed no effects on the replication of the element. Also, no apparent differences in the sensitivity to DNA damaging agents and other stressors or morphological changes in the mutant cells were detected. However, the mutants lacking Ppol showed an increase in two to three orders of magnitude in their transformation efficiency with plasmids and genomic DNA acquired from the environment (eDNA), independently of its origin and G + C content. In contrast, no significant differences with the wild type were detected when the cells received the DNA from other T. thermophilus partners in conjugation-like mating experiments. The similarities of this behaviour with that shown by mutants lacking the Argonaute (ThAgo) protein suggests a putative partnership Ppol-ThAgo in the DNA–DNA interference mechanism of defence, although other eDNA defence mechanisms independent of ThAgo cannot be discarded.post-print697 K

Engineering Permissive Insertion Sites in the Bacteriophage Phi29 DNA-Linked Terminal Protein.

Many different DNA delivery vehicles have been developed and tested, all with their advantages and disadvantages. The bacteriophage phi29 terminal protein (TP) is covalently linked to the 5' ends of the phage genome during the DNA replication process. Our approach is to utilize this TP as a platform to incorporate different protein or peptide modules that can target the DNA to the interior of the cell, to the nucleus, or even to subcellular compartments. In order to be able to insert different peptide modules on the TP sequence to endow it with desired functions and/or eliminate unwanted regions of the protein, we have carried out a transposition screening to detect insertion-permissive points on the sequence of the TP. We report the functional characterization of 12 insertion mutants of the TP, and the identification of one site at position 38 that allows the insertion of peptides up to 17 amino acids in length while maintaining the ability of the TP to support DNA amplification in vitro. A protein with one insertion at that position containing a cysteine residue, a linker, and a thrombin recognition site was purified and its amplification activity was optimized