Pancreatic cancer-initiating cell exosome message transfer into noncancer-initiating cells: the importance of CD44v6 in reprogramming

Background: Cancer-initiating cell (CIC) exosomes (CIC-TEX) are suggested reprogramming Non-CIC. Mode of message transfer and engagement of CIC-markers being disputed, we elaborated the impact of CD44v6 and Tspan8 on the response of Non-CIC. Methods: Non-metastasizing CD44v6- and Tspan8-knockdown (kd) pancreatic cancer cells served as Non-CIC. CIC-TEX coculture-induced changes were evaluated by deep-sequencing and functional assays. Tumor progression was surveyed during in vivo CIC-TEX treatment. Results: Deep-sequencing of CIC-TEX-cocultured CD44v6kd-Non-CIC revealed pronounced mRNA changes in signaling, transport, transcription and translation; altered miRNA affected metabolism, signaling and transcription. CIC-TEX coculture-induced changes in Tspan8kd-Non-CIC mostly relied on CIC-TEX-Tspan8 being required for targeting. CIC-TEX transfer supported apoptosis resistance and significantly promoted epithelial mesenchymal transition, migration, invasion and (lymph)angiogenesis of the kd Non-CIC in vitro and in vivo, deep-sequencing allowing individual mRNA and miRNA assignment to altered functions. Importantly, CIC-TEX act as a hub, initiated by CD44v6-dependent RTK, GPCR and integrin activation and involving CD44v6-assisted transcription and RNA processing. Accordingly, a kinase inhibitor hampered CIC-TEX-fostered tumor progression, which was backed by an anti-Tspan8 blockade of CIC-TEX binding. Conclusions: This in depth report on the in vitro and in vivo impact of CIC-TEX on CD44v6kd and Tspan8kd Non-CIC unravels hub CIC-TEX activity, highlighting a prominent contribution of the CIC-markers CD44v6 to signaling cascade activation, transcription, translation and miRNA processing in Non-CIC and of Tspan8 to CIC-TEX targeting. Blocking CIC-TEX binding/uptake and uptake-initiated target cell activation significantly mitigated the deleterious CIC-TEX impact on CD44v6kd and Tspan8kd Non-CIC

Abnormal P‐wave terminal force in lead V 1 is a marker for atrial electrical dysfunction but not structural remodelling

Aims There is a lack of diagnostic and therapeutic options for patients with atrial cardiomyopathy and paroxysmal atrial fibrillation. Interestingly, an abnormal P-wave terminal force in electrocardiogram lead V1 (PTFV1) has been associated with atrial cardiomyopathy, but this association is poorly understood. We investigated PTFV1 as a marker for functional, electrical, and structural atrial remodelling. Methods and results Fifty-six patients with acute myocardial infarction and 13 kidney donors as control cohort prospectively underwent cardiac magnetic resonance imaging to evaluate the association between PTFV1 and functional remodelling (atrial strain). To further investigate underlying pathomechanisms, right atrial appendage biopsies were collected from 32 patients undergoing elective coronary artery bypass grafting. PTFV1 was assessed as the product of negative P-wave amplitude and duration in lead V1 and defined as abnormal if ≥4000 ms*μV. Activity of cardiac Ca/calmodulin-dependent protein kinase II (CaMKII) was determined by a specific HDAC4 pull-down assay as a surrogate for electrical remodelling. Atrial fibrosis was quantified using Masson's trichrome staining as a measure for structural remodelling. Multivariate regression analyses were performed to account for potential confounders. A total of 16/56 (29%) of patients with acute myocardial infarction, 3/13 (23%) of kidney donors, and 15/32 (47%) of patients undergoing coronary artery bypass grafting showed an abnormal PTFV1. In patients with acute myocardial infarction, left atrial (LA) strain was significantly reduced in the subgroup with an abnormal PTFV1 (LA reservoir strain: 32.28 ± 12.86% vs. 22.75 ± 13.94%, P = 0.018; LA conduit strain: 18.87 ± 10.34% vs. 10.17 ± 8.26%, P = 0.004). Abnormal PTFV1 showed a negative correlation with LA conduit strain independent from clinical covariates (coefficient B: −7.336, 95% confidence interval −13.577 to −1.095, P = 0.022). CaMKII activity was significantly increased from (normalized to CaMKII expression) 0.87 ± 0.17 to 1.46 ± 0.15 in patients with an abnormal PTFV1 (P = 0.047). This increase in patients with an abnormal PTFV1 was independent from clinical covariates (coefficient B: 0.542, 95% confidence interval 0.057 to 1.027, P = 0.031). Atrial fibrosis was significantly lower with 12.32 ± 1.63% in patients with an abnormal PTFV1 (vs. 20.50 ± 2.09%, P = 0.006), suggesting PTFV1 to be a marker for electrical but not structural remodelling. Conclusions Abnormal PTFV1 is an independent predictor for impaired atrial function and for electrical but not for structural remodelling. PTFV1 may be a promising tool to evaluate patients for atrial cardiomyopathy and for risk of atrial fibrillation

Tspan8-Tumor Extracellular Vesicle-Induced Endothelial Cell and Fibroblast Remodeling Relies on the Target Cell-Selective Response

Tumor cell-derived extracellular vesicles (TEX) expressing tetraspanin Tspan8-alpha4/beta1 support angiogenesis. Tspan8-alpha6/beta4 facilitates lung premetastatic niche establishment. TEX-promoted target reprogramming is still being disputed, we explored rat endothelial cell (EC) and lung fibroblast (Fb) mRNA and miRNA profile changes after coculture with TEX. TEX were derived from non-metastatic BSp73AS (AS) or metastatic BSp73ASML (ASML) rat tumor lines transfected with Tspan8 (AS-Tspan8) or Tspan8-shRNA (ASML-Tspan8kd). mRNA was analyzed by deep sequencing and miRNA by array analysis of EC and Fb before and after coculture with TEX. EC and Fb responded more vigorously to AS-Tspan8- than AS-TEX. Though EC and Fb responses differed, both cell lines predominantly responded to membrane receptor activation with upregulation and activation of signaling molecules and transcription factors. Minor TEX-initiated changes in the miRNA profile relied, at least partly, on long noncoding RNA (lncRNA) that also affected chromosome organization and mRNA processing. These analyses uncovered three important points. TEX activate target cell autonomous programs. Responses are initiated by TEX targeting units and are target cell-specific. The strong TEX-promoted lncRNA impact reflects lncRNA shuttling and location-dependent distinct activities. These informations urge for an in depth exploration on the mode of TEX-initiated target cell-specific remodeling including, as a major factor, lncRNA

Long-read only assembly of Drechmeria coniospora genomes reveals widespread chromosome plasticity and illustrates the limitations of current nanopore methods

International audienceBackground: Long-read sequencing is increasingly being used to determine eukaryotic genomes. We used nanopore technology to generate chromosome-level assemblies for 3 different strains of Drechmeria coniospora, a nematophagous fungus used extensively in the study of innate immunity in Caenorhabditis elegans. Results: One natural geographical isolate demonstrated high stability over decades, whereas a second isolate not only had a profoundly altered genome structure but exhibited extensive instability. We conducted an in-depth analysis of sequence errors within the 3 genomes and established that even with state-of-the-art tools, nanopore methods alone are insufficient to generate eukaryotic genome sequences of sufficient accuracy to merit inclusion in public databases. Conclusions: Although nanopore long-read sequencing is not accurate enough to produce publishable eukaryotic genomes, in our case, it has revealed new information about genome plasticity in D. coniospora and provided a backbone that will permit future detailed study to characterize gene evolution in this important model fungal pathogen

Transcriptional analysis of amino acid, metal ion, vitamin and carbohydrate uptake in butanol-producing Clostridium beijerinckii NRRL B-598.

In-depth knowledge of cell metabolism and nutrient uptake mechanisms can lead to the development of a tool for improving acetone-butanol-ethanol (ABE) fermentation performance and help to overcome bottlenecks in the process, such as the high cost of substrates and low production rates. Over 300 genes potentially encoding transport of amino acids, metal ions, vitamins and carbohydrates were identified in the genome of the butanol-producing strain Clostridium beijerinckii NRRL B-598, based on similarity searches in protein function databases. Transcriptomic data of the genes were obtained during ABE fermentation by RNA-Seq experiments and covered acidogenesis, solventogenesis and sporulation. The physiological roles of the selected 81 actively expressed transport genes were established on the basis of their expression profiles at particular stages of ABE fermentation. This article describes how genes encoding the uptake of glucose, iron, riboflavin, glutamine, methionine and other nutrients take part in growth, production and stress responses of C. beijerinckii NRRL B-598. These data increase our knowledge of transport mechanisms in solventogenic Clostridium and may be used in the selection of individual genes for further research