Transcriptomics-based study reveals a novel link between arginine methylation and phosphate regulation in Saccharomyces cerevisiae

Hmt1 is the predominant arginine methyltransferase in yeast, involved in biological processes such as transcription, transcriptional regulation, nucleocytoplasmic transport of proteins and mRNAs and RNA splicing. Hmt1 is also known to mediate protein-protein and protein-RNA interactions. Despite its well-studied role in regulatory processes, the effects of Hmt1 knockout on the transcriptome has never been performed. This thesis investigated the changes in the transcriptome upon deletion of Hmt1, and through this, aimed to uncover as yet undiscovered biological functions of Hmt1. RNA-seq was used to compare the transcriptomic profiles of wild type versus Hmt1-null cells. In mid-log exponential growth, we found dysregulation of phosphate metabolism and polyphosphate accumulation in the Hmt1-null strain. Acid phosphatases (PH05, PH011 and PH012), phosphate transporters (PH084 and PH089) and the vacuolar transporter chaperone VTCJ were amongst genes significantly downregulated in hmt1/J.. Concomitant decreases were observed in intracellular polyphosphate levels and extracellular phosphatase activity. We demonstrated that transcription factor Pho4, responsible for activation of the phosphate regulatory pathway was methylated by Hmt1 in vitro at Arg-241. Genomic mutation of Arg- 241 to Lys in wild type produced to the same phenotype as hmt1/J.. This is the first study to establish a link between arginine methylation and phosphate regulation. As polyphosphate is a source of phosphate, ATP and energy in the yeast cell, we extended our investigation to cells in quiescence. RNA-seq revealed more than 50% genes were differentially expressed between hmt1/J. and wild type quiescent cells. Crucially, phosphate transport was still defective as the membrane-embedded phosphate transporters PH084, PH087, PH089 and PH090, vacuolar polyphosphate transporter PH091 and exopolyphosphatase PPX1 showed significant downregulation in hmt1/J. in quiescence. As a result of this dysregulation in phosphate importation and polyphosphate accumulation in hmt1/J., the Hmt1 deletion strain exhibited advanced quiescence as compared to wild type. A number of models are proposed to explain these observations

IN SILICO METHODS FOR DRUG DESIGN AND DISCOVERY

Computer-aided drug design (CADD) methodologies are playing an ever-increasing role in drug discovery that are critical in the cost-effective identification of promising drug candidates. These computational methods are relevant in limiting the use of animal models in pharmacological research, for aiding the rational design of novel and safe drug candidates, and for repositioning marketed drugs, supporting medicinal chemists and pharmacologists during the drug discovery trajectory.Within this field of research, we launched a Research Topic in Frontiers in Chemistry in March 2019 entitled “In silico Methods for Drug Design and Discovery,” which involved two sections of the journal: Medicinal and Pharmaceutical Chemistry and Theoretical and Computational Chemistry. For the reasons mentioned, this Research Topic attracted the attention of scientists and received a large number of submitted manuscripts. Among them 27 Original Research articles, five Review articles, and two Perspective articles have been published within the Research Topic. The Original Research articles cover most of the topics in CADD, reporting advanced in silico methods in drug discovery, while the Review articles offer a point of view of some computer-driven techniques applied to drug research. Finally, the Perspective articles provide a vision of specific computational approaches with an outlook in the modern era of CADD

Abstracts from the 25th Fungal Genetics Conference

Abstracts from the 25th Fungal Genetics Conferenc

Crystallographic Studies of Enzymes (Volume II)

In this Special Issue of Crystals, entitled "Crystallographic Studies of Enzymes (Volume II)", eleven research papers on key findings and methodologies of structure, function, and reaction mechanisms of enzymes are presented

The Impact of Dynamics in Protein Assembly

Predicting the assembly of multiple proteins into specific complexes is critical to understanding their biological function in an organism, and thus the design of drugs to address their malfunction. Consequently, a significant body of research and development focuses on methods for elucidating protein quaternary structure. In silico techniques are used to propose models that decode experimental data, and independently as a structure prediction tool. These computational methods often consider proteins as rigid structures, yet proteins are inherently flexible molecules, with both local side-chain motion and larger conformational dynamics governing their behaviour. This treatment is particularly problematic for any protein docking engine, where even a simple rearrangement of the side-chain and backbone atoms at the interface of binding partners complicates the successful determination of the correct docked pose. Herein, we present a means of representing protein surface, electrostatics and local dynamics within a single volumetric descriptor, before applying it to a series of physical and biophysical problems to validate it as representative of a protein. We leverage this representation in a protein-protein docking context and demonstrate that its application bypasses the need to compensate for, and predict, specific side-chain packing at the interface of binding partners for both water-soluble and lipid-soluble protein complexes. We find little detriment in the quality of returned predictions with increased flexibility, placing our protein docking approach as highly competitive versus comparative methods. We then explore the role of larger, conformational dynamics in protein quaternary structure prediction, by exploiting large-scale Molecular Dynamics simulations of the SARS-CoV-2 spike glycoprotein to elucidate possible high-order spike-ACE2 oligomeric states. Our results indicate a possible novel path to therapeutics following the COVID-19 pandemic. Overall, we find that the structure of a protein alone is inadequate in understanding its function through its possible binding modes. Therefore, we must also consider the impact of dynamics in protein assembly

Mitochondrial Transport Proteins

Mitochondrial transporters are membrane-inserted proteins which provide a link between metabolic reactions occurring within the mitochondrial matrix and outside the organelles by catalyzing the translocation of numerous solutes across the mitochondrial membrane. They include the mitochondrial carrier family members, the proteins involved in pyruvate transport, ABC transporters and channels, and are, therefore, essential for many biological processes and cell homeostasis. Identification and functional studies of many mitochondrial transporters have been performed over the years using both in vitro and in vivo systems. The few recently solved structures of these transporters have paved the way for further investigations. Furthermore, alterations in their function are responsible for several diseases

Role of Protein-Protein Interactions in Metabolism: Genetics, Structure, Function

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