Cofitness network connectivity determines a fuzzy essential zone in open bacterial pangenome

Most in silico evolutionary studies commonly assumed that core genes are essential for cellular function,while accessory genes are dispensable,particularly in nutrient-rich environments.However,this assumption is seldom tested genetically within the pangenome context.In this study,we conducted a robust pangenomic Tn-seq analysis of fitness genes in a nutrient-rich medium for Sinorhizobium strains with a canonical open pangenome.To evaluate the robustness of fitness category assignment,Tn-seq data for three independent mutant libraries per strain were analyzed by three methods,which indicates that the Hidden Markov Model(HMM)-based method is most robust to variations between mutant libraries and not sensitive to data size,outperforming the Bayesian and Monte Carlo simulation-based methods.Consequently,the HMM method was used to classify the fitness category.Fitness genes,categorized as essential(ES),advantage(GA),and disadvantage(GD)genes for growth,are enriched in core genes,while nonessential genes(NE)are over-represented in accessory genes.Accessory ES/GA genes showed a lower fitness effect than core ES/GA genes.Connectivity degrees in the cofitness network decrease in the order of ES,GD,and GA/NE.In addition to accessory genes,1599 out of 3284 core genes display differential essentiality across test strains.Within the pangenome core,both shared quasi-essential(ES and GA)and strain-dependent fitness genes are enriched in similar functional categories.Our analysis demonstrates a considerable fuzzy essential zone determined by cofitness connectivity degrees in Sinorhizobium pangenome and highlights the power of the cofitness network in understanding the genetic basis of ever-increasing prokaryotic pangenome data.

RNA therapeutics:updates and future potential

Recent advancements in the production,modification,and cellular delivery of RNA molecules facilitated the expansion of RNAbased therapeutics.The increasing understanding of RNA biology initiated a corresponding growth in RNA therapeutics.In this review,the general concepts of five classes of RNA-based therapeutics,including RNA interference-based therapies,antisense oligonucleotides,small activating RNA therapies,circular RNA therapies,and messenger RNA-based therapeutics,will be discussed.Moreover,we also provide an overview of RNA-based therapeutics that have already received regulatory approval or are currently being evaluated in clinical trials,along with challenges faced by these technologies.RNA-based drugs demonstrated positive clinical trial results and have the ability to address previously“undruggable”targets,which delivers great promise as a disruptive therapeutic technology to fulfill its full clinical potentiality.

RNAi screen to identify protein phosphatases that regulate the NF-kappaB signaling

NF-kappaB plays a critical role in cell survival,apoptosis,and inflammatory responses.Serine/threoninespecific phosphatases(PPs)represent the second major class of enzymes that catalyze the dephosphorylation of proteins.The roles of PPs regulating NF-kappaB activities are poorly understood.Here we describe an RNAi-based screen to identify the PPs that involve in regulating NFkappaB signaling.Thirty-four candidate PPs siRNAs were synthesized and primarily screened by NF-kappaB reporter gene assay in HeLa cells.PHLPP,one of the protein phosphatase type 2C family members(PP2C),was identified as a positive regulator of NF-kappaB signaling.Knock-down of PHLPP dramatically attenuated TNFα-stimulated NF-kappaB transcriptional activation.Knockdown of PHLPP led to enhancement of NF-kappaB/p65 nuclear import and retention,but decreased TNFα-induced phosphorylation at Ser276 on p65.This critical phosphorylation was also drastically reduced by knock-down of PKCalpha and Akt1,two important serine/threonine kinases dephosphorylated by PHLPP.The results together suggest that PHLPP-Akt-PKC may represent an important signaling loop that activates NF-kappaB/p65 signaling through critical serine phosphorylation.