Current antiretroviral therapy (ART) provides potent suppression of HIV-1 replication. However, ART does not target latent viral reservoirs, so persistent infection remains a challenge. Small molecules with pharmacological properties that allow them to reach and activate reservoirs could potentially be utilized eliminate the arm when used in combination ART. Here we describe cell-based system modeling latency was high-throughput screen identify small molecule antagonists latency. A more...

Here, we present a modification to single-molecule fluorescence in situ hybridization that enables quantitative detection and analysis of small RNA (sRNA) expressed bacteria. We show short (∼200 nucleotide) nucleic acid targets can be detected when the background unbound singly dye-labeled DNA oligomers is reduced through with set complementary labeled quencher. By neutralizing from probes, were able significantly reduce number false positives, allowing for accurate quantification sRNA...

Abstract Background Human induced pluripotent stem cells (iPSC) have opened new avenues for regenerative medicine. Consequently, iPSC-derived motor neurons emerged as potentially viable therapies spinal cord injuries and neurodegenerative disorders including Amyotrophic Lateral Sclerosis. However, direct clinical application of iPSC bears in itself the risk tumorigenesis other unforeseeable genetic or epigenetic abnormalities. Results Employing RNA-seq technology, we identified characterized...

Bacterial small RNAs (sRNAs) regulate gene expression by base-pairing with downstream target mRNAs to attenuate translation of mRNA into protein at the post-transcriptional level. In response specific environmental changes, sRNAs can modulate levels genes, thus enabling adaptation cellular physiology. We profiled sRNA in Gram-negative bacteria Burkholderia thailandensis cultured under 54 distinct growth conditions using a Burkholderia-specific microarray that contains probe sets all...

Burkholderia pseudomallei is the causative agent of melioidosis, a disease with mortality rate up to 40% even treatment. Despite ability certain antibiotics control initial infection, relapse occurs in treated patients. The inability clear this bacterial infection part due persistence, an evasion mechanism against and effect host defenses. Evaluation antibiotic efficacy B. revealed that 48% vitro grown populations can survive persister state. Toxin–antitoxin (TA) systems have been previously...

Human immunodeficiency virus type 1 (HIV-1) is not eliminated from patients even after years of antiretroviral therapy, apparently due to the presence latently infected cells. Here we describe development a cell-based system latency that can be used for high-throughput screening aimed at novel drug discovery eradicate HIV-1 infection.

<title>Abstract</title> Chromatin conformation capture followed by next-generation sequencing in combination with large-scale polymer simulations (4DHiC) produces detailed information on genomic loci interactions, allowing for the interrogation of 3D spatial structures. Here, Hi-C data was acquired from infection fetal lung fibroblast (MRC5) cells α-coronavirus 229E (CoV229E). Experimental contact maps were used to determine viral-induced changes architecture over a 48-hour time period...

Non-coding small RNAs (sRNAs) are found in practically all bacterial genomes and play important roles regulating gene expression to impact metabolism, growth, virulence. We performed transcriptomics analysis identify sRNAs that differentially expressed Yersinia pestis invaded the human macrophage cell line THP-1, compared pathogens remained extracellular presence of host. Using ultra high-throughput sequencing, we identified 37 novel 143 previously known Y. pestis. In particular, sRNA Ysr170...

Abstract Background The pathogenic Yersinia species exhibit a primarily extracellular lifestyle through manipulation of host signaling pathways that regulate pro-inflammatory gene expression and cytokine release. To identify genes are targeted by during the infection process, we performed an RNA interference (RNAi) screen based on recovery NF-κB-mediated activation in response to TNF-α stimulation upon Y. enterocolitica infection. Results We screened shRNAs against 782 human kinome 26 heat...

Interactions between the cytoplasmic domains of viral transmembrane proteins and host machinery often determine outcome infection. The M2 protein influenza A has been identified as a key player in autophagy-mediated replication. Here, we describe engineering validation an antibody specific for domain protein. Through phage yeast display selection techniques, obtained that recognizes: 1) purified from bacterial inclusion bodies refolded, 2) full-length recombinant expressed mammalian cells,...

Bacterial persistence, known as noninherited antibacterial resistance, is a factor contributing to the establishment of long-lasting chronic bacterial infections. In this study, we examined ability nicotinamide (NA) potentiate activity different classes antibiotics against Burkholderia thailandensis persister cells.

Persister cells are genetically identical variants in a bacterial population that have phenotypically modified their physiology to survive environmental stress. In pathogens, persisters able antibiotic treatment and reinfect patients frustrating cycle of chronic infection. To better define core persistence mechanisms for therapeutics development, we performed transcriptomics analyses Burkholderia thailandensis populations enriched via three methods: flow sorting low proton motive force,...

Neuromuscular junctions (NMJs), specialized synapses between motor neurons and muscle fibers, are essential for activity. A simple reproducible cell-based in vitro NMJ platform is needed to test the impact of chemicals on neuron-muscle communication. Our utilizes genetically modified cells, optimized culture conditions, commercially available multielectrode array system recording action potentials. Neuronal cells (NSC34) were optogenetically with channelrhodopsin chimera allow simultaneous,...

Abstract Autophagy is a critical mechanism deployed by eukaryotic cells in response to stress, including viral infection, boost the innate antimicrobial responses. However, an increasing number of pathogens hijack autophagic machinery facilitate their own replication. Influenza A virus (IAV), responsible for several global pandemics, has intricate dependence on autophagy successful replication mammalian cells. To elucidate key chokepoints host stress responses facilitating IAV replication,...

Neurodegeneration causes a significant disease burden and there are few therapeutic interventions available for reversing or slowing the progression. Induced pluripotent stem cells (iPSCs) hold potential since they sourced from adult tissue have capacity to be differentiated into numerous cell lineages, including motor neurons. This differentiation process traditionally relies on lineage patterning factors supplied in media. Genetic engineering of iPSC with introduction recombinant master...

Pathogenic Burkholderia rely on host factors for efficient intracellular replication and are highly refractory to antibiotic treatment. To identify genes that required by spp during infection, we performed a RNA interference (RNAi) screen of the human kinome identified 35 kinases facilitated thailandensis survival in monocytic THP-1 cells. We validated selection using imaging flow cytometry assess efficiency B. upon siRNA-mediated knockdown. focused role novel protein kinase C isoform,...

Persistence is a bet-hedging strategy in bacterial populations that increases antibiotic tolerance and leads to the establishment of latent infections. In this study, we demonstrated synthetic non-toxic taxane-based reversal agent (tRA), developed as an inhibitor ABC transporter systems mammalian cancer cells, enhanced killing persister from different pathogens, including Burkholderia, Pseudomonas, Francisella, Yersinia. Acting efflux at 100 nM, tRA99020 efficiency suppressed production...