The acetate uptake transporter family motif "NPAPLGL(M/S)" is essential for substrate uptake

Organic acids are recognized as one of the most prevalent compounds in ecosystems, thus the transport and assimilation of these molecules represent an adaptive advantage for organisms. The AceTr family members are associated with the active transport of organic acids, namely acetate and succinate. The phylogenetic analysis shows this family is dispersed in the tree of life. However, in eukaryotes, it is almost limited to microbes, though reaching a prevalence close to 100% in fungi, with an essential role in spore development. Aiming at deepening the knowledge in this family, we studied the acetate permease AceP from Methanosarcina acetivorans, as the first functionally characterized archaeal member of this family. Furthermore, we demonstrate that the yeast Gpr1 from Yarrowia lipolytica is an acetate permease, whereas the Ady2 closest homologue in Saccharomyces cerevisiae, Fun34, has no role in acetate uptake. In this work, we describe the functional role of the AceTr conserved motif NPAPLGL(M/S). We further unveiled the role of the amino acid residues R122 and Q125 of SatP as essential for protein activity.This work was supported by the strategic program UID/BIA/04050/2013 (POCI-01-0145-FEDER-007569) and the project PTDC/BIAMIC/5184/2014 funded by national funds through the Fundacao para a Ciencia e Tecnologia (FCT) I.P. and by the European Regional Development Fund (ERDF) through the COMPETE 2020 - Programa Operacional Competitividade e Internacionalizacao (POCI); by the project EcoAgriFood: Innovative green products and processes to promote AgriFood BioEconomy (operacao NORTE-01-0145-FEDER-000009), supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). Work at ITQB NOVA was financially supported by Project LISBOA-01-0145-FEDER-007660 (Microbiologia Molecular, Estrutural e Celular) funded by FEDER funds through COMPETE2020 - Programa Operacional Competitividade e Intemacionalizacao (POCI). DR acknowledges FCT for the SFRH/BD/96166/2013 PhD grant. MSS acknowledges the Norte2020 for the UMINHO/BD/25/2016 PhD grant with the reference NORTE-08-5369-FSE-000060

Meeting report – Cell dynamics: host–pathogen interface

Two years into the most significant infectious disease event of our generation, infections have populated every conversation and in-depth understanding of host-pathogen interactions has, perhaps, never been more important. In a successful return to in-person conferences, the host-pathogen interface was the focus of the third Cell Dynamics meeting, which took place at the glorious Wotton House in Surrey, UK. The meeting organised by Michaela Gack, Maximiliano Gutierrez, Dominique Soldati-Favre and Michael Way gathered an international group of scientists who shared their recent discoveries and views on numerous aspects, including cell-autonomous defence mechanisms, pathogen interactions with host cytoskeletal or membrane dynamics, and cellular immune regulation. More than 30 years into the beginning of cellular microbiology as a field, the meeting exhibited the unique aspect of the host-pathogen interface in uncovering the fundamentals of both pathogens and their hosts

Exploring the function and structure of the acetate transporter family

The AceTr transporter family is a group of Acetate Transporters with six predicted transmembrane segments. It has homologues in fungi, bacteria, archaea and protozoa. As acetate transporters, these proteins play a crucial role on cell metabolism being involved in the capacity of cells to adapt to nutrient availability. As these membrane transporters also transport other substrates, they may be an important asset for biotechnological purposes. To better understand the mechanism of acetate transport, we have investigated functional and structural features of the AceTr family. We have performed site-directed mutagenesis in residues highly conserved, or possibly involved in substrate binding, from the Saccharomyces cerevisiae Ady2. Functional assays determined the influence of these residues on the transport capacity and protein localization. Another goal of this work is to determine the structure of the AceTr family. Accordingly, several of its members were cloned in different expression vectors and their expression was evaluated in different E. coli expression strains. SatP from E. coli and its AceP from Methanosarcina acetivorans presented satisfactory levels of protein production and are currently being tested in production, purification and crystallization trials

Following Darwin's footsteps: Evaluating the impact of an activity designed for elementary school students to link historically important evolution key concepts on their understanding of natural selection

While several researchers have suggested that evolution should be explored from the initial years of schooling, little information is available on effective resources to enhance elementary school students' level of understanding of evolution by natural selection (LUENS). For the present study, we designed, implemented, and evaluated an educational activity planned for fourth graders (9 to 10 years old) to explore concepts and conceptual fields that were historically important for the discovery of natural selection. Observation field notes and students' productions were used to analyze how the students explored the proposed activity. Additionally, an evaluation framework consisting of a test, the evaluation criteria, and the scoring process was applied in two fourth-grade classes (N = 44) to estimate elementary school students' LUENS before and after engaging in the activity. Our results show that our activity allowed students to link the key concepts, resulting in a significant increase of their understanding of natural selection. They also reveal that additional activities and minor fine-tuning of the present activity are required to further support students' learning about the concept of differential reproduction

Klebsiella pneumoniae type VI secretion system-mediated microbial competition is PhoPQ controlled and reactive oxygen species dependent

Copyright: © 2020 Storey et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Klebsiella pneumoniae is recognized as an urgent threat to human health due to the increasing isolation of multidrug resistant strains. Hypervirulent strains are a major concern due to their ability to cause life-threating infections in healthy hosts. The type VI secretion system (T6SS) is widely implicated in microbial antagonism, and it mediates interactions with host eukaryotic cells in some cases. In silico search for genes orthologous to T6SS component genes and T6SS effector genes across 700 K. pneumoniae genomes shows extensive diversity in T6SS genes across the K. pneumoniae species. Temperature, oxygen tension, pH, osmolarity, iron levels, and NaCl regulate the expression of the T6SS encoded by a hypervirulent K. pneumoniae strain. Polymyxins and human defensin 3 also increase the activity of the T6SS. A screen for regulators governing T6SS uncover the correlation between the transcription of the T6SS and the ability to kill E. coli prey. Whereas H-NS represses the T6SS, PhoPQ, PmrAB, Hfq, Fur, RpoS and RpoN positively regulate the T6SS. K. pneumoniae T6SS mediates intra and inter species bacterial competition. This antagonism is only evident when the prey possesses an active T6SS. The PhoPQ two component system governs the activation of K. pneumoniae T6SS in bacterial competitions. Mechanistically, PhoQ periplasmic domain, and the acid patch within, is essential to activate K. pneumoniae T6SS. Klebsiella T6SS also mediates anti-fungal competition. We have delineated the contribution of each of the individual VgrGs in microbial competition and identified VgrG4 as a T6SS effector. The DUF2345 domain of VgrG4 is sufficient to intoxicate bacteria and yeast. ROS generation mediates the antibacterial effects of VgrG4, and the antitoxin Sel1E protects against the toxic activity of VgrG4. Our findings provide a better understanding of the regulation of the T6SS in bacterial competitions, and place ROS as an early event in microbial competition