RNase2 is the member of the RNaseA family most abundant in macrophages. Here, we knocked out RNase2 in THP-1 cells and analysed the response to Respiratory Syncytial Virus (RSV). RSV induced RNase2 expression, which significantly enhanced cell survival. Next, by cP-RNAseq sequencing, which amplifies the cyclic-phosphate endonuclease products, we analysed the ncRNA population. Among the ncRNAs accumulated in WT vs KO cells, we found mostly tRNA-derived fragments (tRFs) and second miRNAs. Differential sequence coverage identified tRFs from only few parental tRNAs, revealing a predominant cleavage at anticodon and d-loops at U/C (B1) and A (B2) sites. Selective tRNA cleavage was confirmed in vitro using the recombinant protein. Likewise, only few miRNAs were significantly more abundant in WT vs RNase2-KO cells. Complementarily, by screening of a tRF & tiRNA array, we identified an enriched population associated to RNase2 expression and RSV exposure. The results confirm the protein antiviral action and provide the first evidence of its cleavage selectivity on ncRNAs. Graphical abstract

Tissue plasminogen activator (tPA) is an effective treatment for ischemic stroke, but its neurotoxicity is a significant problem. Here we tested the hypothesis that recombinant ADAMTS 13 (rADAMTS 13) would reduce tPA neurotoxicity in a mouse model of stroke. We show that treatment with rADAMTS 13 in combination with tPA significantly reduced infarct volume compared with mice treated with tPA alone 48 hours after stroke. The combination treatment significantly improved neurological deficits compared with mice treated with tPA or vehicle alone. These neuroprotective effects were associated with significant reductions in fibrin deposits in ischemic vessels and less severe cell death in ischemic brain. The effect of rADAMTS13 on tPA neurotoxicity was mimicked by the N-methyl-D-aspartate (NMDA) receptor antagonist M-801 and was abolished by injection of NMDA. Moreover, rADAMTS 13 prevents the neurotoxicity effect of tPA, by blocking its interaction with the NMDA receptor NR2B and the attendant phosphorylation of NR2B and activation of ERK1/2. Finally, the NR2B-specific NMDA receptor antagonist ifenprodil abolished tPA neurotoxicity and rADAMTS 13 treatment had no further beneficial effect. Our data suggest that the combination of rADAMTS 13 and tPA may provide a novel treatment of ischemic stroke by diminishing the neurotoxic effects of exogenous tPA.

Liver development is a complex process that is regulated by a series of signaling pathways. Three-dimensional (3D) chromatin architecture plays an important role in transcriptional regulation; nonetheless, its dynamics and role in the rapid transition of core liver functions during development and obesity-induced metabolic stress remain largely unexplored. To investigate the dynamic chromatin architecture during liver development and under metabolic stress, we generated high-resolution maps of chromatin architecture for porcine livers across six major developmental stages (from embryonic day 38 to the adult stage) and under a high-fat diet-induced obesity. The characteristically loose chromatin architecture supports a highly plastic genome organization during early liver development, which fundamentally contributes to the rapid functional transitions in the liver after birth. We reveal the multi-scale reorganization of chromatin architecture and its influence on transcriptional regulation of critical signaling processes during liver development, and show its close association with transition in hepatic functions (i.e., from hematopoiesis in the fetus to metabolism and immunity after birth). The limited changes in chromatin structure help explain the observed metabolic adaptation to excessive energy intake in pigs. These results provide a global overview of chromatin architecture dynamics associated with the transition of physiological liver functions between prenatal development and postnatal maturation, and a foundational resource that allows for future in-depth functional characterization.

The human RNase3 is a member of the RNaseA superfamily involved in host immunity. RNase3 is expressed by leukocytes and shows broad-spectrum antimicrobial activity. Together with a direct antimicrobial action, RNase3 exhibits immunomodulatory properties. Here, we have analysed the transcriptome of macrophages exposed to the wild-type protein and a catalytic-defective mutant (RNase3-H15A). The analysis of differently expressed genes (DEGs) in treated THP1-derived macrophages highlighted a common pro-inflammatory “core-response” independent of the protein ribonucleolytic activity. Network analysis identified the epidermal growth factor receptor (EGFR) as the main central regulatory protein. Expression of selected DEGs and MAPK phosphorylation were inhibited by an anti-EGFR antibody. Structural analysis suggested that RNase3 activates the EGFR pathway by direct interaction with the receptor. Besides, we identified a subset of DEGs related to the protein ribonucleolytic activity, characteristic of virus infection response. Transcriptome analysis revealed an early pro-inflammatory response, not associated to the protein catalytic activity, followed by a late activation in a ribonucleolytic-dependent manner. Next, we demonstrated that overexpression of macrophage endogenous RNase3 protects the cells against infection byMycobacterium aurumand the human respiratory syncytial virus. Comparison of cell infection profiles in the presence of Erlotinib, an EGFR inhibitor, revealed that the receptor activation is required for the antibacterial but not for the antiviral protein action. Moreover, the DEGs related and unrelated to the protein catalytic activity are associated to the immune response to bacterial and viral infection, respectively. We conclude that RNase3 modulates the macrophage defence against infection in both catalytic-dependent and independent manners.