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Institute
As matchmaker between mRNA and sRNA interactions, the RNA chaperone Hfq plays a key role in riboregulation of many bacteria. Often, the global influence of Hfq on the transcriptome is reflected by substantially altered proteomes and pleiotropic phenotypes in hfq mutants. Using quantitative proteomics and co-immunoprecipitation combined with RNA-sequencing (RIP-seq) of Hfq-bound RNAs, we demonstrate the pervasive role of Hfq in nutrient acquisition, metabolism and motility of the plant pathogen Agrobacterium tumefaciens. 136 of 2544 proteins identified by iTRAQ (isobaric tags for relative and absolute quantitation) were affected in the absence of Hfq. Most of them were associated with ABC transporters, general metabolism and motility. RIP-seq of chromosomally encoded Hfq 3xFlag revealed 1697 mRNAs and 209 non-coding RNAs (ncRNAs) associated with Hfq. 56 ncRNAs were previously undescribed. Interestingly, 55% of the Hfq-bound ncRNAs were encoded antisense (as) to a protein-coding sequence suggesting that A. tumefaciens Hfq plays an important role in asRNA-target interactions. The exclusive enrichment of 296 mRNAs and 31 ncRNAs under virulence conditions further indicates a role for post-transcriptional regulation in A. tumefaciens-mediated plant infection. On the basis of the iTRAQ and RIP-seq data, we assembled a comprehensive model of the Hfq core regulon in A. tumefaciens.
A shear-dependent NO-cGMP-cGKI cascade in platelets acts as an auto-regulatory brake of thrombosis
(2018)
Mechanisms that limit thrombosis are poorly defined. One of the few known endogenous platelet inhibitors is nitric oxide (NO). NO activates NO sensitive guanylyl cyclase (NO-GC) in platelets, resulting in an increase of cyclic guanosine monophosphate (cGMP). Here we show, using cGMP sensor mice to study spatiotemporal dynamics of platelet cGMP, that NO-induced cGMP production in pre-activated platelets is strongly shear-dependent. We delineate a new mode of platelet-inhibitory mechanotransduction via shear-activated NO-GC followed by cGMP synthesis, activation of cGMP-dependent protein kinase I (cGKI), and suppression of Ca2+ signaling. Correlative profiling of cGMP dynamics and thrombus formation in vivo indicates that high cGMP concentrations in shear-exposed platelets at the thrombus periphery limit thrombosis, primarily through facilitation of thrombus dissolution. We propose that an increase in shear stress during thrombus growth activates the NO-cGMP-cGKI pathway, which acts as an auto-regulatory brake to prevent vessel occlusion, while preserving wound closure under low shear.