Iron-Utilization System in Vibrio vulnificus M2799

Vibrio vulnificus is a Gram-negative pathogenic bacterium that causes serious infections in humans and requires iron for growth. A clinical isolate, V. vulnificus M2799, secretes a catecholate siderophore, vulnibactin, that captures ferric ions from the environment. In the ferric-utilization system in V. vulnificus M2799, an isochorismate synthase (ICS) and an outer membrane receptor, VuuA, are required under low-iron conditions, but alternative proteins FatB and VuuB can function as a periplasmic-binding protein and a ferric-chelate reductase, respectively. The vulnibactin-export system is assembled from TolCV1 and several RND proteins, including VV1_1681. In heme acquisition, HupA and HvtA serve as specific outer membrane receptors and HupB is a sole periplasmic-binding protein, unlike FatB in the ferric-vulnibactin utilization system. We propose that ferric-siderophore periplasmic-binding proteins and ferric-chelate reductases are potential targets for drug discovery in infectious diseases.

Machine Learning Uncovers a Data-Driven Transcriptional Regulatory Network for the Crenarchaeal Thermoacidophile Sulfolobus acidocaldarius

Dynamic cellular responses to environmental constraints are coordinated by the transcriptional regulatory network (TRN), which modulates gene expression. This network controls most fundamental cellular responses, including metabolism, motility, and stress responses. Here, we apply independent component analysis, an unsupervised machine learning approach, to 95 high-quality Sulfolobus acidocaldarius RNA-seq datasets and extract 45 independently modulated gene sets, or iModulons. Together, these iModulons contain 755 genes (32% of the genes identified on the genome) and explain over 70% of the variance in the expression compendium. We show that five modules represent the effects of known transcriptional regulators, and hypothesize that most of the remaining modules represent the effects of uncharacterized regulators. Further analysis of these gene sets results in: (1) the prediction of a DNA export system composed of five uncharacterized genes, (2) expansion of the LysM regulon, and (3) evidence for an as-yet-undiscovered global regulon. Our approach allows for a mechanistic, systems-level elucidation of an extremophile’s responses to biological perturbations, which could inform research on gene-regulator interactions and facilitate regulator discovery in S. acidocaldarius. We also provide the first global TRN for S. acidocaldarius. Collectively, these results provide a roadmap toward regulatory network discovery in archaea.

Machine learning uncovers a data-driven transcriptional regulatory network for the Crenarchaeal thermoacidophile Sulfolobus acidocaldarius

Dynamic cellular responses to environmental constraints are coordinated by the transcriptional regulatory network (TRN), which modulates gene expression. This network controls most fundamental cellular responses, including metabolism, motility, and stress responses. Here, we apply independent component analysis, an unsupervised machine learning approach, to 95 high-quality Sulfolobus acidocaldarius RNA-seq datasets and extract 45 independently modulated gene sets, or iModulons. Together, these iModulons contain 755 genes (32% of the genes identified on the genome) and explain over 70% of the variance in the expression compendium. We show that 5 modules represent the effects of known transcriptional regulators, and hypothesize that most of the remaining modules represent the effects of uncharacterized regulators. Further analysis of these gene sets results in: (1) the prediction of a DNA export system composed of 5 uncharacterized genes, (2) expansion of the LysM regulon, and (3) evidence for an as-yet-undiscovered global regulon. Our approach allows for a mechanistic, systems-level elucidation of an extremophile's responses to biological perturbations, which could inform research on gene-regulator interactions and facilitate regulator discovery in S. acidocaldarius. We also provide the first global TRN for S. acidocaldarius. Collectively, these results provide a roadmap towards regulatory network discovery in archaea.

Development of Experimental Prototypes for the Teaching of Science

The following document presents the development of educational experimental prototypes. The themes of the prototypes cover principles or concepts of physics. Like kinematics, oscillatory movements and shocks. Each prototype has sensors that allow measurements of the variables involved in the experimentation. At the same time, mobile applications are developed for devices with Android operating system, which allow the user to control the prototype from their cell phone and also view the variables measured by the experimentation prototypes. The applications also have a data export system that allows to obtain a file in csv format. In this way, the measurements are obtained in the form of a table to later be processed through office automation tools or other numerical computing software such as Matlab.

Vesicular and uncoated Rab1-dependent cargo carriers facilitate ER to Golgi transport

ABSTRACTSecretory cargo is recognized, concentrated and trafficked from endoplasmic reticulum (ER) exit sites (ERES) to the Golgi. Cargo export from the ER begins when a series of highly conserved COPII coat proteins accumulate at the ER and regulate the formation of cargo-loaded COPII vesicles. In animal cells, capturing live de novo cargo trafficking past this point is challenging; it has been difficult to discriminate whether cargo is trafficked to the Golgi in a COPII-coated vesicle. Here, we describe a recently developed live-cell cargo export system that can be synchronously released from ERES to illustrate de novo trafficking in animal cells. We found that components of the COPII coat remain associated with the ERES while cargo is extruded into COPII-uncoated, non-ER associated, Rab1 (herein referring to Rab1a or Rab1b)-dependent carriers. Our data suggest that, in animal cells, COPII coat components remain stably associated with the ER at exit sites to generate a specialized compartment, but once cargo is sorted and organized, Rab1 labels these export carriers and facilitates efficient forward trafficking.This article has an associated First Person interview with the first author of the paper.

Rules of Expansion: an Updated Consensus Operator Site for the CopR-CopY Family of Bacterial Copper Exporter System Repressors

ABSTRACT Copper is broadly toxic to bacteria. As such, bacteria have evolved specialized copper export systems (cop operons) often consisting of a DNA-binding/copper-responsive regulator (which can be a repressor or activator), a copper chaperone, and a copper exporter. For those bacteria using DNA-binding copper repressors, few studies have examined the regulation of this operon regarding the operator DNA sequence needed for repressor binding. In Streptococcus pneumoniae (the pneumococcus), CopY is the copper repressor for the cop operon. Previously, homologs of pneumococcal CopY have been characterized to bind a 10-base consensus sequence T/GACANNTGTA known as the cop box. Using this motif, we sought to determine whether genes outside the cop operon are also regulated by the CopY repressor, which was previously shown in Lactococcus lactis. We found that S. pneumoniae CopY did not bind to cop operators upstream of these candidate genes in vitro. During this process, we found that the cop box sequence is necessary but not sufficient for CopY binding. Here, we propose an updated operator sequence for the S. pneumoniae cop operon to be ATTGACAAATGTAGAT binding CopY with a dissociation constant (Kd) of ∼28 nM. We demonstrate strong cross-species interaction between some CopY proteins and CopY operators, suggesting strong evolutionary conservation. Taken together with our binding studies and bioinformatics data, we propose the consensus operator RNYKACANNYGTMRNY for the bacterial CopR-CopY copper repressor homologs. IMPORTANCE Many Gram-positive bacteria respond to copper stress by upregulating a copper export system controlled by a copper-sensitive repressor, CopR-CopY. The previous operator sequence for this family of proteins had been identified as TACANNTGTA. Here, using several recombinant proteins and mutations in various DNA fragments, we define those 10 bases as necessary but not sufficient for binding and in doing so, refine the cop operon operator to the 16-base sequence RNYKACANNTGTMRNY. Due to the sheer number of repressors that have been said to bind to the original 10 bases, including many antibiotic resistance repressors such as BlaI and MecI, we feel that this study highlights the need to reexamine many of these sites of the past and use added stringency for verifying operators in the future.

A Novel Efficient L-Lysine Exporter Identified by Functional Metagenomics

Lack of active export system often limits the industrial bio-based production processes accumulating the intracellular product and hence complexing the purification steps. L-lysine, an essential amino acid, is produced biologically in quantities exceeding two million tons per year; yet, L-lysine production is challenged by efficient export system at high titres during fermentation. To address this issue, new exporter candidates for efficient efflux of L-lysine are needed. Using metagenomic functional selection, we identified 58 genes encoded on 28 unique metagenomic fragments from cow gut microbiome library that improved L-lysine tolerance. These genes include a novel putative L-lysine transporter, belonging to a previously uncharacterized EamA superfamily. Characterization using Xenopus oocyte expression system as well as an Escherichia coli host demonstrates activity as a L-lysine transporter. This novel exporter improved L-lysine tolerance in E. coli by 40% and enhanced the specific productivity of L-lysine in an industrial Corynebacterium glutamicum strain by 12%. Our approach allows the sequence-independent discovery of novel exporters and can be deployed to increase titres and productivity of toxicity-limited bioprocesses.

Determination of factors involved in the rejection of bananas (Musa acuminata) intended for international commercialization

Rendering production and trade more sustainable can be accomplished via promoting innovation and sustainable business models. This paper addresses sustainable production and international trade in the banana agro-export sector of Ecuador. This activity provides millions of dollars in income, but with this development, a series of quality standards have been established to enter the competitive export system. These criteria contributed to establishing good post-harvest production and management practices that guarantee optimal banana and plantain production. The objective of this study is to determine the factors involved in the rejection of bananas (Musa acuminata) intended for international commercialization. Our methodology considered the design modality of non-experimental transactional research, using a quantitative approach. Methodological design was developed in three phases at Finca 6 Hermanas, located in the Barraganete sector of the San Juan parish in the Puebloviejo canton of the Los Ríos Province, Ecuador. Results highlight that 79.55 % of reasons for banana rejection are abiotic factors (damage, dry latex, scar, insect damage, broken neck, and overgrowth), while biotic factors ( twins, diseases, and short finger) cause 20.45%. Over the 6-week duration of our investigation the average rejection was 6,361 fingers and 1,269 kg. The analysis of variance turned out to be significant for variable 1 (biotic and abiotic). In this case the null hypothesis (Ho) is rejected; with the criterion of p-value < 0.0001 and F (9; 45) = 2.10., F = 13.17> F critical. For variable (2) “work weeks”, Ho is accepted for p-value of 0.7694 and F (5; 45) = 2.4. As F = 0.51 < F critical, one can conclude, that with a significance level of 5% the null hypothesis is accepted. It is also established that these figures enable strategies that systemically mitigate the damages via correcting the causes that lead to the deterioration of banana and, by this, increase the economic gains of commercialization.