Genomic and phenotypic insight into Xanthomonas vesicatoria strains with different aggressiveness on tomato.

Xanthomonas vesicatoria is one of the causal agents of bacterial spot, a disease that seriously affects the production of tomato (Solanum lycopersicum) and pepper (Capsicum annum) worldwide. In Argentina, bacterial spot is found in all tomato producing areas, with X. vesicatoria being one of the main species detected in the fields. Previously, we isolated three X. vesicatoria strains BNM 208, BNM 214, and BNM 216 from tomato plants with bacterial spot, and found they differed in their ability to form biofilm and in their degree of aggressiveness. Here, the likely causes of those differences were explored through genotypic and phenotypic studies. The genomes of the three strains were sequenced and assembled, and then compared with each other and also with 12 other publicly available X. vesicatoria genomes. Phenotypic characteristics (mainly linked to biofilm formation and virulence) were studied in vitro. Our results show that the differences observed earlier between BNM 208, BNM 214, and BNM 216 may be related to the structural characteristics of the xanthan gum produced by each strain, their repertoire of type III effectors (T3Es), the presence of certain genes associated with c-di-GMP metabolism and type IV pili (T4P). These findings on the pathogenicity mechanisms of X. vesicatoria could be useful for developing bacterial spot control strategies aimed at interfering with the infection processes.

Analysis of changes of cavity volumes in predefined directions of protein motions and cavity flexibility.

Dynamics of protein cavities associated with protein fluctuations and conformational plasticity is essential for their biological function. NMR ensembles, molecular dynamics (MD) simulations, and normal mode analysis (NMA) provide appropriate frameworks to explore functionally relevant protein dynamics and cavity changes relationships. Within this context, we have recently developed analysis of null areas (ANA), an efficient method to calculate cavity volumes. ANA is based on a combination of algorithms that guarantees its robustness against numerical differentiations. This is a unique feature with respect to other methods. Herein, we present an updated and improved version that expands it use to quantify changes in cavity features, like volume and flexibility, due to protein structural distortions performed on predefined biologically relevant directions, for example, directions of largest contribution to protein fluctuations (principal component analysis [PCA modes]) obtained by MD simulations or ensembles of NMR structures, collective NMA modes or any other direction of motion associated with specific conformational changes. A web page has been developed where its facilities are explained in detail. First, we show that ANA can be useful to explore gradual changes of cavity volume and flexibility associated with protein ligand binding. Secondly, we perform a comparison study of the extent of variability between protein backbone structural distortions, and changes in cavity volumes and flexibilities evaluated for an ensemble of NMR active and inactive conformers of the epidermal growth factor receptor structures. Finally, we compare changes in size and flexibility between sets of NMR structures for different homologous chains of dynein.

PED in 2021: a major update of the protein ensemble database for intrinsically disordered proteins.

The Protein Ensemble Database (PED) (https://proteinensemble.org), which holds structural ensembles of intrinsically disordered proteins (IDPs), has been significantly updated and upgraded since its last release in 2016. The new version, PED 4.0, has been completely redesigned and reimplemented with cutting-edge technology and now holds about six times more data (162 versus 24 entries and 242 versus 60 structural ensembles) and a broader representation of state of the art ensemble generation methods than the previous version. The database has a completely renewed graphical interface with an interactive feature viewer for region-based annotations, and provides a series of descriptors of the qualitative and quantitative properties of the ensembles. High quality of the data is guaranteed by a new submission process, which combines both automatic and manual evaluation steps. A team of biocurators integrate structured metadata describing the ensemble generation methodology, experimental constraints and conditions. A new search engine allows the user to build advanced queries and search all entry fields including cross-references to IDP-related resources such as DisProt, MobiDB, BMRB and SASBDB. We expect that the renewed PED will be useful for researchers interested in the atomic-level understanding of IDP function, and promote the rational, structure-based design of IDP-targeting drugs.

Exploring Conformational Space with Thermal Fluctuations Obtained by Normal-Mode Analysis.

Proteins in their native states can be represented as ensembles of conformers in dynamical equilibrium. Thermal fluctuations are responsible for transitions between these conformers. Normal-modes analysis (NMA) using elastic network models (ENMs) provides an efficient procedure to explore global dynamics of proteins commonly associated with conformational transitions. In the present work, we present an iterative approach to explore protein conformational spaces by introducing structural distortions according to their equilibrium dynamics at room temperature. The approach can be used either to perform unbiased explorations of conformational space or to explore guided pathways connecting two different conformations, e.g., apo and holo forms. In order to test its performance, four proteins with different magnitudes of structural distortions upon ligand binding have been tested. In all cases, the conformational selection model has been confirmed and the conformational space between apo and holo forms has been encompassed. Different strategies have been tested that impact on the efficiency either to achieve a desired conformational change or to achieve a balanced exploration of the protein conformational multiplicity.

Network analysis of dynamically important residues in protein structures mediating ligand-binding conformational changes.

According to the generalized conformational selection model, ligand binding involves the co-existence of at least two conformers with different ligand-affinities in a dynamical equilibrium. Conformational transitions between them should be guaranteed by intramolecular vibrational dynamics associated to each conformation. These motions are, therefore, related to the biological function of a protein. Positions whose mutations are found to alter these vibrations the most can be defined as key positions, that is, dynamically important residues that mediate the ligand-binding conformational change. In a previous study, we have shown that these positions are evolutionarily conserved. They correspond to buried aliphatic residues mostly localized in regular structured regions of the protein like ß-sheets and α-helices. In the present paper, we perform a network analysis of these key positions for a large dataset of paired protein structures in the ligand-free and ligand-bound form. We observe that networks of interactions between these key positions present larger and more integrated networks with faster transmission of the information. Besides, networks of residues result that are robust to conformational changes. Our results reveal that the conformational diversity of proteins seems to be guaranteed by a network of strongly interconnected key positions rather than individual residues.

The effect of winter length on duration of dormancy and survival of specialized herbivorous Rhagoletis fruit flies from high elevation environments with acyclic climatic variability.

Dormancy can be defined as a state of suppressed development allowing insects to cope with adverse conditions and plant phenology. Among specialized herbivorous insects exploiting seasonal resources, diapause frequently evolves as a strategy to adjust to predictable plant seasonal cycles. To cope with acyclic and unpredictable climatic events, it has been found for some insects that a proportion of the population undergoes prolonged dormancy. We compared the response of three species in the Rhagoletis cingulata species group exploiting plants differing in fruiting phenology from environments varying in frequency and timing of acyclic climatic catastrophic events (frost during flowering and fruit set) and varying also in the time of the onset of the rainy season. Small proportions (10 months), and large proportions of pupae died without emerging as adults. The number of days elapsed from the end of artificial winter and adult eclosion was longer for R. cingulata exploiting late fruiting Prunus serotina in Northeastern Mexico than for flies recovered from earlier fruiting plants in the central Altiplano. Rhagoletis turpiniae and northeastern R. cingulata pupae suffered high proportions of parasitism. Large proportions of R. cingulata from central Mexico engaging in prolonged dormancy may be explained by the fact that flowering and fruit set for its host, P. serotina var capuli, driven by the timing of maximum precipitation, matches a period of highest probability of frost often resulting in large areas with fruitless trees at unpredictable time intervals. As a consequence of differences in host plant fruiting phenology, central and northeastern Mexican R. cingulata were found to be allochronically isolated. Prolonged dormancy may have resulted in escape from parasitism.

Structural and dynamics evidence for scaffold asymmetric flexibility of the human transthyretin tetramer.

The molecular symmetry of multimeric proteins is generally determined by using X-ray diffraction techniques, so that the basic question as to whether this symmetry is perfectly preserved for the same protein in solution remains open. In this work, human transthyretin (TTR), a homotetrameric plasma transport protein with two binding sites for the thyroid hormone thyroxine (T4), is considered as a case study. Based on the crystal structure of the TTR tetramer, a hypothetical D2 symmetry is inferred for the protein in solution, whose functional behavior reveals the presence of two markedly different Kd values for the two T4 binding sites. The latter property has been ascribed to an as yet uncharacterized negative binding cooperativity. A triple mutant form of human TTR (F87M/L110M/S117E TTR), which is monomeric in solution, crystallizes as a tetrameric protein and its structure has been determined. The exam of this and several other crystal forms of human TTR suggests that the TTR scaffold possesses a significant structural flexibility. In addition, TTR tetramer dynamics simulated using normal modes analysis exposes asymmetric vibrational patterns on both dimers and thermal fluctuations reveal small differences in size and flexibility for ligand cavities at each dimer-dimer interface. Such small structural differences between monomers can lead to significant functional differences on the TTR tetramer dynamics, a feature that may explain the functional heterogeneity of the T4 binding sites, which is partially overshadowed by the crystal state.

Development of a Low-Cost and Effective Trapping Device for Apple Maggot Fly (Diptera: Tephritidae) Monitoring and Control in Mexican Commercial Hawthorn Groves.

Few efforts have been made in Mexico to monitor Rhagoletis pomonella (Walsh) (Diptera: Tephritidae) in commercial hawthorn (Crataegus spp.) crops. Therefore, the main objectives of this study were to evaluate infestation levels of R. pomonella in feral and commercial Mexican hawthorn and to assess the efficacy of different trap-lure combinations to monitor the pest. Wild hawthorn was more infested than commercially grown hawthorn at the sample site. No differences among four commercial baits (Biolure, ammonium carbonate, CeraTrap, and Captor + borax) were detected when used in combination with a yellow sticky gel (SG) adherent trap under field conditions. However, liquid lures elicited a slightly higher, although not statistically different, capture. Cage experiments in the laboratory revealed that flies tended to land more often on the upper and middle than lower-bottom part of polyethylene (PET) bottle traps with color circles. Among red, orange, green, and yellow circles attached to a bottle trap, only yellow circles improved fly captures compared with a colorless trap. A PET bottle trap with a red circle over a yellow background captured more flies than a similar trap with yellow circles. An SG adherent yellow panel trap baited with ammonium carbonate was superior to the improved PET bottle trap (red over a yellow background) baited with different liquid proteins, but a higher proportion of females and no differences in fly detection were measured in PET traps baited with protein lures. These trials open the door for future research into development of a conventional nonadherent trap to monitor or control R. pomonella.

Evaluating the effect of mutations and ligand binding on transthyretin homotetramer dynamics.

Native transthyretin (TTR) homotetramer dissociation is the first step of the fibrils formation process in amyloid disease. A large number of specific point mutations that destabilize TTR quaternary structure have shown pro-amyloidogenic effects. Besides, several compounds have been proposed as drugs in the therapy of TTR amyloidosis due to their TTR tetramer binding affinities, and therefore, contribution to its integrity. In the present paper we have explored key positions sustaining TTR tetramer dynamical stability. We have identified positions whose mutations alter the most the TTR tetramer equilibrium dynamics based on normal mode analysis and their response to local perturbations. We have found that these positions are mostly localized at ß-strands E and F and EF-loop. The monomer-monomer interface is pointed out as one of the most vulnerable regions to mutations that lead to significant changes in the TTR-tetramer equilibrium dynamics and, therefore, induces TTR amyloidosis. Besides, we have found that mutations on residues localized at the dimer-dimer interface and/or at the T4 hormone binding site destabilize the tetramer more than the average. Finally, we were able to compare several compounds according to their effect on vibrations associated to the ligand binding. Our ligand comparison is discussed and analyzed in terms of parameters and measurements associated to TTR-ligand binding affinities and the stabilization of its native state.

Conformational diversity analysis reveals three functional mechanisms in proteins.

Protein motions are a key feature to understand biological function. Recently, a large-scale analysis of protein conformational diversity showed a positively skewed distribution with a peak at 0.5 Å C-alpha root-mean-square-deviation (RMSD). To understand this distribution in terms of structure-function relationships, we studied a well curated and large dataset of ~5,000 proteins with experimentally determined conformational diversity. We searched for global behaviour patterns studying how structure-based features change among the available conformer population for each protein. This procedure allowed us to describe the RMSD distribution in terms of three main protein classes sharing given properties. The largest of these protein subsets (~60%), which we call "rigid" (average RMSD = 0.83 Å), has no disordered regions, shows low conformational diversity, the largest tunnels and smaller and buried cavities. The two additional subsets contain disordered regions, but with differential sequence composition and behaviour. Partially disordered proteins have on average 67% of their conformers with disordered regions, average RMSD = 1.1 Å, the highest number of hinges and the longest disordered regions. In contrast, malleable proteins have on average only 25% of disordered conformers and average RMSD = 1.3 Å, flexible cavities affected in size by the presence of disordered regions and show the highest diversity of cognate ligands. Proteins in each set are mostly non-homologous to each other, share no given fold class, nor functional similarity but do share features derived from their conformer population. These shared features could represent conformational mechanisms related with biological functions.

Transcriptional Network Architecture of Breast Cancer Molecular Subtypes.

Breast cancer heterogeneity is evident at the clinical, histological and molecular level. High throughput technologies allowed the identification of intrinsic subtypes that capture transcriptional differences among tumors. A remaining question is whether said differences are associated to a particular transcriptional program which involves different connections between the same molecules. In other words, whether particular transcriptional network architectures can be linked to specific phenotypes. In this work we infer, construct and analyze transcriptional networks from whole-genome gene expression microarrays, by using an information theory approach. We use 493 samples of primary breast cancer tissue classified in four molecular subtypes: Luminal A, Luminal B, Basal and HER2-enriched. For comparison, a network for non-tumoral mammary tissue (61 samples) is also inferred and analyzed. Transcriptional networks present particular architectures in each breast cancer subtype as well as in the non-tumor breast tissue. We find substantial differences between the non-tumor network and those networks inferred from cancer samples, in both structure and gene composition. More importantly, we find specific network architectural features associated to each breast cancer subtype. Based on breast cancer networks' centrality, we identify genes previously associated to the disease, either, generally (i.e., CNR2) or to a particular subtype (such as LCK). Similarly, we identify LUZP4, a gene barely explored in breast cancer, playing a role in transcriptional networks with subtype-specific relevance. With this approach we observe architectural differences between cancer and non-cancer at network level, as well as differences between cancer subtype networks which might be associated with breast cancer heterogeneity. The centrality measures of these networks allow us to identify genes with potential biomedical implications to breast cancer.

The effect of winter length on survival and duration of dormancy of four sympatric species of Rhagoletis exploiting plants with different fruiting phenology.

Dormancy has been thoroughly studied for several species of economic importance in the genus Rhagoletis in temperate areas of North America and Europe. Much less is known on life history regulation for species inhabiting high-elevation areas in the subtropics at the southern extreme of their geographical range. Host plant phenology has been found to play a key role in generating allochronic isolation among sibling species and host races of Rhagoletis in the course of sympatric speciation, and has important implications for pest management. We compare the effect of winter length on survival to adult eclosion and dormancy duration among four species of Rhagoletis (three of them sympatric) exploiting hosts with different fruiting phenology in subtropical isolated highlands. Survival and duration of dormancy was found to be different among the four species. At 24°C, a very small proportion (<1%) of R. pomonella, R. turpiniae and R. zoqui completed development without becoming dormant, while in the case of R. solanophaga the majority of the population emerged after development within 40 days of pupation. Also, a large proportion of braconid parasitoids infesting Rhagoletis eggs and larvae emerged as adults without becoming dormant. Greatest survival after artificial winter was obtained for R. pomonella (50-60%) and R. zoqui (30%) after only four weeks at 5°C (a third of the time reported for studies on northern R. pomonella), while R. turpiniae, under identical environmental conditions experienced low adult emergence, and highest survival (11%) was recorded for flies exposed to 5°C during 10 and 12 weeks. For R. pomonella, there was a strong positive relationship between winter length and time to post-winter adult eclosion that was not observed for R. zoqui. In sum, for R. pomonella, mild winters in highland subtropical areas appear to select for flies better able to withstand longer periods of warm temperature before winter than flies exploiting late fruiting hosts and inhabiting northern latitudes. In the case of R. turpiniae and R. zoqui environmental cues such as fluctuations in humidity and/or different temperature thresholds (5°C) may play a more important role than winter length in life history regulation. Continuous host availability for R. solanophaga appears to have selected for non-diapausing flies. From an applied perspective our results are useful for handling flies in the laboratory to conduct research and suggest that non-diapausing strains of flies and parasitoids may be selected for SIT and innundative biological control programs.

Community Structure Reveals Biologically Functional Modules in MEF2C Transcriptional Regulatory Network.

Gene regulatory networks are useful to understand the activity behind the complex mechanisms in transcriptional regulation. A main goal in contemporary biology is using such networks to understand the systemic regulation of gene expression. In this work, we carried out a systematic study of a transcriptional regulatory network derived from a comprehensive selection of all potential transcription factor interactions downstream from MEF2C, a human transcription factor master regulator. By analyzing the connectivity structure of such network, we were able to find different biologically functional processes and specific biochemical pathways statistically enriched in communities of genes into the network, such processes are related to cell signaling, cell cycle and metabolism. In this way we further support the hypothesis that structural properties of biological networks encode an important part of their functional behavior in eukaryotic cells.

Evolutionary Conserved Positions Define Protein Conformational Diversity.

Conformational diversity of the native state plays a central role in modulating protein function. The selection paradigm sustains that different ligands shift the conformational equilibrium through their binding to highest-affinity conformers. Intramolecular vibrational dynamics associated to each conformation should guarantee conformational transitions, which due to its importance, could possibly be associated with evolutionary conserved traits. Normal mode analysis, based on a coarse-grained model of the protein, can provide the required information to explore these features. Herein, we present a novel procedure to identify key positions sustaining the conformational diversity associated to ligand binding. The method is applied to an adequate refined dataset of 188 paired protein structures in their bound and unbound forms. Firstly, normal modes most involved in the conformational change are selected according to their corresponding overlap with structural distortions introduced by ligand binding. The subspace defined by these modes is used to analyze the effect of simulated point mutations on preserving the conformational diversity of the protein. We find a negative correlation between the effects of mutations on these normal mode subspaces associated to ligand-binding and position-specific evolutionary conservations obtained from multiple sequence-structure alignments. Positions whose mutations are found to alter the most these subspaces are defined as key positions, that is, dynamically important residues that mediate the ligand-binding conformational change. These positions are shown to be evolutionary conserved, mostly buried aliphatic residues localized in regular structural regions of the protein like ß-sheets and α-helix.

A home telehealth program for patients with severe COPD: the PROMETE study.

BACKGROUND: Acute exacerbations of chronic obstructive pulmonary disease (AECOP) are key events in the natural history of the disease. Patients with more AECOPD have worse prognosis. There is a need of innovative models of care for patients with severe COPD and frequent AECOPD, and Telehealth (TH) is part of these programs. METHODS: In a cluster assignment, controlled trial study design, we recruited 60 patients, 30 in home telehealth (HT) and 30 in conventional care (CC). All participants had a prior diagnosis of COPD with a post-bronchodilator forced expiratory volume (FEV1)% predicted <50%, age ≥ 50 years, were on long-term home oxygen therapy, and non-smokers. Patients in the HT group measured their vital signs on a daily bases, and data were transmitted automatically to a Clinical Monitoring Center for followed-up, and who escalated clinical alerts to a Pneumologist. RESULTS: After 7-month of monitoring and follow-up, there was a significant reduction in ER visits (20 in HT vs. 57 in CC), hospitalizations (12 vs. 33), length of hospital stay in (105 vs. 276 days), and even need for non-invasive mechanical ventilation (0 vs. 8), all p < 0.05. Time to the first severe AECOPD increased from 77 days in CC to 141 days in HT (K-M p < 0.05). There was no study withdrawals associated with technology. All patients showed a high level of satisfaction with the HT program. CONCLUSIONS: We conclude that HT in elderly, severe COPD patients with multiple comorbidities is safe and efficacious in reducing healthcare resources utilization.

Isolation and genotype analysis of rubella virus from a case of Guillain-Barré syndrome.

Rubella virus (RV) infection has sporadically been linked to Guillain-Barré syndrome (GBS), but the association with RV has been based only on clinical and/or serological backgrounds. In the present case it was possible to isolate RV (genotype 1a) from cerebrospinal fluid and peripheral blood mononuclear cells of an 18-year-old woman diagnosed with GBS after clinical manifestations of rubella. This report contributes to confirm RV as one of the triggering pathogens of this peripheral nervous system disease.

Educación, salud y estreñimiento / Education, health and constipation

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