A comprehensive SARS-CoV-2 and COVID-19 review, Part 2: host extracellular to systemic effects of SARS-CoV-2 infection.

COVID-19, the disease caused by SARS-CoV-2, has caused significant morbidity and mortality worldwide. The betacoronavirus continues to evolve with global health implications as we race to learn more to curb its transmission, evolution, and sequelae. The focus of this review, the second of a three-part series, is on the biological effects of the SARS-CoV-2 virus on post-acute disease in the context of tissue and organ adaptations and damage. We highlight the current knowledge and describe how virological, animal, and clinical studies have shed light on the mechanisms driving the varied clinical diagnoses and observations of COVID-19 patients. Moreover, we describe how investigations into SARS-CoV-2 effects have informed the understanding of viral pathogenesis and provide innovative pathways for future research on the mechanisms of viral diseases.

RNA Regulatory Networks 2.0.

The central role of RNA molecules in cell biology has been an expanding subject of study since the proposal of the "RNA world" hypothesis 60 years ago [...].

Transcriptomic Analysis of Acetaminophen Biodegradation by Penicillium chrysogenum var. halophenolicum and Insights into Energy and Stress Response Pathways.

(1) Background: Acetaminophen (APAP), an active component of many analgesic and antipyretic drugs, is one of the most concerning trace contaminants in the environment and is considered as an emergent pollutant of marine and aquatic ecosystems. Despite its biodegradability, APAP has become a recalcitrant compound due to the growth of the global population, the ease of availability, and the inefficient wastewater treatment applied. (2) Methods: In this study, we used a transcriptomic approach to obtain functional and metabolic insights about the metabolization of APAP by a phenol-degrading fungal strain, Penicillium chrysogenum var. halophenolicum. (3) Results: We determined that the transcriptomic profile exhibited by the fungal strain during APAP degradation was very dynamic, being characterized by an abundance of dysregulated transcripts which were proportional to the drug metabolization. Using a systems biology approach, we also inferred the protein functional interaction networks that could be related to APAP degradation. We proposed the involvement of intracellular and extracellular enzymes, such as amidases, cytochrome P450, laccases, and extradiol-dioxygenases, among others. (4) Conclusions: Our data suggested that the fungus could metabolize APAP via a complex metabolic pathway, generating nontoxic metabolites, which demonstrated its potential in the bioremediation of this drug.

The interplay between lncRNAs, RNA-binding proteins and viral genome during SARS-CoV-2 infection reveals strong connections with regulatory events involved in RNA metabolism and immune response.

Rationale: Viral infections are complex processes based on an intricate network of molecular interactions. The infectious agent hijacks components of the cellular machinery for its profit, circumventing the natural defense mechanisms triggered by the infected cell. The successful completion of the replicative viral cycle within a cell depends on the function of viral components versus the cellular defenses. Non-coding RNAs (ncRNAs) are important cellular modulators, either promoting or preventing the progression of viral infections. Among these ncRNAs, the long non-coding RNA (lncRNA) family is especially relevant due to their intrinsic functional properties and ubiquitous biological roles. Specific lncRNAs have been recently characterized as modulators of the cellular response during infection of human host cells by single stranded RNA viruses. However, the role of host lncRNAs in the infection by human RNA coronaviruses such as SARS-CoV-2 remains uncharacterized. Methods: In the present work, we have performed a transcriptomic study of a cohort of patients with different SARS-CoV-2 viral load and analyzed the involvement of lncRNAs in supporting regulatory networks based on their interaction with RNA-binding proteins (RBPs). Results: Our results revealed the existence of a SARS-CoV-2 infection-dependent pattern of transcriptional up-regulation in which specific lncRNAs are an integral component. To determine the role of these lncRNAs, we performed a functional correlation analysis complemented with the study of the validated interactions between lncRNAs and RBPs. This combination of in silico functional association studies and experimental evidence allowed us to identify a lncRNA signature composed of six elements - NRIR, BISPR, MIR155HG, FMR1-IT1, USP30-AS1, and U62317.2 - associated with the regulation of SARS-CoV-2 infection. Conclusions: We propose a competition mechanism between the viral RNA genome and the regulatory lncRNAs in the sequestering of specific RBPs that modulates the interferon response and the regulation of RNA surveillance by nonsense-mediated decay (NMD).

A Structural View of miRNA Biogenesis and Function.

Micro-RNAs (miRNAs) are a class of non-coding RNAs (ncRNAs) that act as post-transcriptional regulators of gene expression. Since their discovery in 1993, they have been the subject of deep study due to their involvement in many important biological processes. Compared with other ncRNAs, miRNAs are generated from devoted transcriptional units which are processed by a specific set of endonucleases. The contribution of structural biology methods for understanding miRNA biogenesis and function has been essential for the dissection of their roles in cell biology and human disease. In this review, we summarize the application of structural biology for the characterization of the molecular players involved in miRNA biogenesis (processors and effectors), starting from the X-ray crystallography methods to the more recent cryo-electron microscopy protocols.

Systematic Structure-Based Search for Ochratoxin-Degrading Enzymes in Proteomes from Filamentous Fungi.

(1) Background: ochratoxins are mycotoxins produced by filamentous fungi with important implications in the food manufacturing industry due to their toxicity. Decontamination by specific ochratoxin-degrading enzymes has become an interesting alternative for the treatment of contaminated food commodities. (2) Methods: using a structure-based approach based on homology modeling, blind molecular docking of substrates and characterization of low-frequency protein motions, we performed a proteome mining in filamentous fungi to characterize new enzymes with potential ochratoxinase activity. (3) Results: the proteome mining results demonstrated the ubiquitous presence of fungal binuclear zinc-dependent amido-hydrolases with a high degree of structural homology to the already characterized ochratoxinase from Aspergillus niger. Ochratoxinase-like enzymes from ochratoxin-producing fungi showed more favorable substrate-binding pockets to accommodate ochratoxins A and B. (4) Conclusions: filamentous fungi are an interesting and rich source of hydrolases potentially capable of degrading ochratoxins, and could be used for the detoxification of diverse food commodities.

Non-Random Genome Editing and Natural Cellular Engineering in Cognition-Based Evolution.

Neo-Darwinism presumes that biological variation is a product of random genetic replication errors and natural selection. Cognition-Based Evolution (CBE) asserts a comprehensive alternative approach to phenotypic variation and the generation of biological novelty. In CBE, evolutionary variation is the product of natural cellular engineering that permits purposive genetic adjustments as cellular problem-solving. CBE upholds that the cornerstone of biology is the intelligent measuring cell. Since all biological information that is available to cells is ambiguous, multicellularity arises from the cellular requirement to maximize the validity of available environmental information. This is best accomplished through collective measurement purposed towards maintaining and optimizing individual cellular states of homeorhesis as dynamic flux that sustains cellular equipoise. The collective action of the multicellular measurement and assessment of information and its collaborative communication is natural cellular engineering. Its yield is linked cellular ecologies and mutualized niche constructions that comprise biofilms and holobionts. In this context, biological variation is the product of collective differential assessment of ambiguous environmental cues by networking intelligent cells. Such concerted action is enabled by non-random natural genomic editing in response to epigenetic impacts and environmental stresses. Random genetic activity can be either constrained or deployed as a 'harnessing of stochasticity'. Therefore, genes are cellular tools. Selection filters cellular solutions to environmental stresses to assure continuous cellular-organismal-environmental complementarity. Since all multicellular eukaryotes are holobionts as vast assemblages of participants of each of the three cellular domains (Prokaryota, Archaea, Eukaryota) and the virome, multicellular variation is necessarily a product of co-engineering among them.

Structural Insights into Carboxylic Polyester-Degrading Enzymes and Their Functional Depolymerizing Neighbors.

Esters are organic compounds widely represented in cellular structures and metabolism, originated by the condensation of organic acids and alcohols. Esterification reactions are also used by chemical industries for the production of synthetic plastic polymers. Polyester plastics are an increasing source of environmental pollution due to their intrinsic stability and limited recycling efforts. Bioremediation of polyesters based on the use of specific microbial enzymes is an interesting alternative to the current methods for the valorization of used plastics. Microbial esterases are promising catalysts for the biodegradation of polyesters that can be engineered to improve their biochemical properties. In this work, we analyzed the structure-activity relationships in microbial esterases, with special focus on the recently described plastic-degrading enzymes isolated from marine microorganisms and their structural homologs. Our analysis, based on structure-alignment, molecular docking, coevolution of amino acids and surface electrostatics determined the specific characteristics of some polyester hydrolases that could be related with their efficiency in the degradation of aromatic polyesters, such as phthalates.

Interspecies Communication in Holobionts by Non-Coding RNA Exchange.

Complex organisms are associations of different cells that coexist and collaborate creating a living consortium, the holobiont. The relationships between the holobiont members are essential for proper homeostasis of the organisms, and they are founded on the establishment of complex inter-connections between all the cells. Non-coding RNAs are regulatory molecules that can also act as communication signals between cells, being involved in either homeostasis or dysbiosis of the holobionts. Eukaryotic and prokaryotic cells can transmit signals via non-coding RNAs while using specific extracellular conveyors that travel to the target cell and can be translated into a regulatory response by dedicated molecular machinery. Within holobionts, non-coding RNA regulatory signaling is involved in symbiotic and pathogenic relationships among the cells. This review analyzes current knowledge regarding the role of non-coding RNAs in cell-to-cell communication, with a special focus on the signaling between cells in multi-organism consortia.

Si vis pacem para bellum: A prospective in silico analysis of miRNA-based plant defenses against fungal infections.

Fungal pathogens are an important threat for plant crops, being responsible for important reductions of production yields and a consequent economic impact. Among the molecular mediators of fungal infections of plant crops, non-coding RNAs (ncRNAs) have been described as relevant players either in the plant immune responses and mechanism of defense or in the colonization of plant tissues by fungi. Acting as a mechanism of defense, some plant small ncRNAs such as miRNAs and tasiRNAs can be secreted by cells and directed to target the transcriptome of pathogenic fungi, triggering an RNAi-like interference mechanism able to silence the expression of specific fungal genes. The detailed knowledge of this mechanism of defense against fungal pathogens could open new possibilities for the protection of human important crops. To infer putative functional relationships mediated by ncRNA communication, we performed a prospective analysis to determine potential plant miRNAs able to target the genome of fungal pathogens, which resulted in the description of enriched specific plant miRNA families and their putative fungal targets that could be further studied in the context of plant-fungi interactions. The expression profile of specific members of the enriched miRNAs families showed an infection-dependent behavior in laboratory models of infection. Plant miRNAs showed sequence complementarity with coding genes of their cognate fungal pathogens. Plant miRNAs could potentially target fungal genes belonging to functional families related to stress response, membrane architecture, vacuolar transport, membrane traffic, and anabolic processes. Families of specific infection-responsive miRNAs are included in the putative plant defense mechanism.

Simultaneous removal of dihydroxybenzenes and toxicity reduction by Penicillium chrysogenum var. halophenolicum under saline conditions.

The dihydroxybenzenes are widely found in wastewater and usually more than one of these aromatic compounds co-exist as pollutants of water resources. The current study investigated and compared the removal efficiency of hydroquinone, catechol and resorcinol in binary substrate systems under saline conditions by Penicillium chrysogenum var. halophenolicum, to clarify the potential of this fungal strain to degrade these aromatic compounds. Since P. chrysogenum is a known penicillin producer, biosynthetic penicillin genes were examined and antibiotic was quantified in mono and binary dihydroxybenzene systems to elucidate the carbon flux of dihydroxybenzenes metabolism in the P. chrysogenum var. halophenolicum to the secondary metabolism. In binary substrate systems, the three assayed dihydroxybenzene compounds were found to be co-metabolized by fungal strain. The fungal strain preferentially degraded hydroquinone and catechol. Resorcinol was degraded slower and supports higher antibiotic titers than either catechol or hydroquinone. Dihydroxybenzenes were faster removed in mixtures compared to mono substrate systems, except for the case of hydroquinone. In this context, the expression of penicillin biosynthetic gene cluster was not related to the removal of dihydroxybenzenes. Penicillin production was triggered simultaneously or after dihydroxybenzene degradation, but penicillin yields, under these conditions, did not compromise dihydroxybenzene biological treatment. To investigate the decrease in dihydroxybenzenes toxicity due to the fungal activity, viability tests with human colon cancer cells (HCT116) and DNA damage by alkaline comet assays were performed. For all the conditions assays, a decrease in saline medium toxicity was observed, indicating its potential as detoxification agent.

Applications of genome editing by programmable nucleases to the metabolic engineering of secondary metabolites.

Genome engineering is a branch of modern biotechnology composed of a cohort of protocols designed to construct and modify a genotype with the main objective of giving rise to a desired phenotype. Conceptually, genome engineering is based on the so called genome editing technologies, a group of genetic techniques that allow either to delete or to insert genetic information in a particular genomic locus. Ten years ago, genome editing tools were limited to virus-driven integration and homologous DNA recombination. However, nowadays the uprising of programmable nucleases is rapidly changing this paradigm. There are two main families of modern tools for genome editing depending on the molecule that controls the specificity of the system and drives the editor machinery to its place of action. Enzymes such as Zn-finger and TALEN nucleases are protein-driven genome editors; while CRISPR system is a nucleic acid-guided editing system. Genome editing techniques are still not widely applied for the design of new compounds with pharmacological activity, but they are starting to be considered as promising tools for rational genome manipulation in biotechnology applications. In this review we will discuss the potential applications of programmable nucleases for the metabolic engineering of secondary metabolites with biological activity.

Gibberellins in Penicillium strains: Challenges for endophyte-plant host interactions under salinity stress.

The genus Penicillium is one of the most versatile "mycofactories", comprising some species able to produce gibberellins, bioactive compounds that can modulate plant growth and development. Although plants have the ability to synthesize gibberellins, their levels are lower when plants are under salinity stress. It has been recognized that detrimental abiotic conditions, such as saline stress, have negative effects on plants, being the availability of bioactive gibberellins a critical factor for their growth under this conditions. This review summarizes the interplay existing between endophytic Penicillium strains and plant host interactions, with focus on bioactive gibberellins production as a fungal response that allows plants to overcome salinity stress.

Noncoding Transcriptional Landscape in Human Aging.

Aging is a universal phenomenon in metazoans, characterized by a general decline of the organism physiology associated with an increased risk of mortality and morbidity. Aging of an organism correlates with a decline in function of its cells, as shown for muscle, immune, and neuronal cells. As the DNA content of most cells within an organism remains largely identical throughout the life span, age-associated transcriptional changes must be achieved by epigenetic mechanisms. However, how aging may impact on the epigenetic state of cells is only beginning to be understood. In light of a growing number of studies demonstrating that noncoding RNAs can provide molecular signals that regulate expression of protein-coding genes and define epigenetic states of cells, we hypothesize that noncoding RNAs could play a direct role in inducing age-associated profiles of gene expression. In this context, the role of long noncoding RNAs (lncRNAs) as regulators of gene expression might be important for the overall transcriptional landscape observed in aged human cells. The possible functions of lncRNAs and other noncoding RNAs, and their roles in the regulation of aging-related cellular pathways will be analyzed.

Transcriptomic Crosstalk between Fungal Invasive Pathogens and Their Host Cells: Opportunities and Challenges for Next-Generation Sequencing Methods.

Fungal invasive infections are an increasing health problem. The intrinsic complexity of pathogenic fungi and the unmet clinical need for new and more effective treatments requires a detailed knowledge of the infection process. During infection, fungal pathogens are able to trigger a specific transcriptional program in their host cells. The detailed knowledge of this transcriptional program will allow for a better understanding of the infection process and consequently will help in the future design of more efficient therapeutic strategies. Simultaneous transcriptomic studies of pathogen and host by high-throughput sequencing (dual RNA-seq) is an unbiased protocol to understand the intricate regulatory networks underlying the infectious process. This protocol is starting to be applied to the study of the interactions between fungal pathogens and their hosts. To date, our knowledge of the molecular basis of infection for fungal pathogens is still very limited, and the putative role of regulatory players such as non-coding RNAs or epigenetic factors remains elusive. The wider application of high-throughput transcriptomics in the near future will help to understand the fungal mechanisms for colonization and survival, as well as to characterize the molecular responses of the host cell against a fungal infection.

Effects of Penicillium chrysogenum var. halophenolicum on kraft lignin: color stabilization and cytotoxicity evaluation.

Wood industries and agricultural crops generate an inexhaustible supply of by-products like lignin, which constitutes an environmental problem. Increasing efforts have been done to find new applications for lignin. One of them is as a food additive, but its chemical nature makes it sensitive to browning which constitutes a major drawback for this type of lignin application. In the present study we are documenting how color stabilization of a commercial kraft lignin was achieved after the treatment with Penicillium chrysogenum var. halophenolicum. In addition the fungal capacity to remove lignin is studied together with the effect of its treatment on cytotoxicity of lignin. P. chrysogenum var. halophenolicum was able to transform lignin, ensuring its color stability for more than 24 months. Dynamic light scattering and atomic force microscopy showed that the fungus contributed to homogenize particle size and hydrodynamic properties in lignin suspensions without increase the toxicity over HeLa cells and human primary fibroblasts. These findings suggest new uses for kraft lignin after P. chrysogenum var. halophenolicum treatment providing an effective approach for improve color stability.

Unzippers, resolvers and sensors: a structural and functional biochemistry tale of RNA helicases.

The centrality of RNA within the biological world is an irrefutable fact that currently attracts increasing attention from the scientific community. The panoply of functional RNAs requires the existence of specific biological caretakers, RNA helicases, devoted to maintain the proper folding of those molecules, resolving unstable structures. However, evolution has taken advantage of the specific position and characteristics of RNA helicases to develop new functions for these proteins, which are at the interface of the basic processes for transference of information from DNA to proteins. RNA helicases are involved in many biologically relevant processes, not only as RNA chaperones, but also as signal transducers, scaffolds of molecular complexes, and regulatory elements. Structural biology studies during the last decade, founded in X-ray crystallography, have characterized in detail several RNA-helicases. This comprehensive review summarizes the structural knowledge accumulated in the last two decades within this family of proteins, with special emphasis on the structure-function relationships of the most widely-studied families of RNA helicases: the DEAD-box, RIG-I-like and viral NS3 classes.

MicroRNA profiling in plasma or serum using quantitative RT-PCR.

MicroRNAs (miRNAs) are important cellular modulators that regulate gene expression at the posttranscriptional level. Circulating miRNAs present in human plasma or serum have recently become an emerging field in biomedical research, mostly due to its potential applications in the diagnosis and prognosis of several diseases. Although miRNA profiling in biofluids holds great promise, there are several challenges to overcome. Here, we present an experimental procedure for profiling miRNA in plasma or serum with high sensitivity and specificity using qRT-PCR. This method is also suitable for studying miRNAs in other body fluids or clinical samples that also contain low amounts of RNA.