cGAS recruitment to micronuclei is dictated by pre-existing nuclear chromatin status.

Micronuclei (MN) are aberrant cytosolic compartments containing broken genomic fragments or whole lagging chromosomes. MN envelopes irreversibly rupture, allowing the viral receptor cGAS to localize to MN and initiate an inflammatory signalling cascade. Here, we demonstrate that MN envelope rupture is not sufficient for cGAS localization. Unlike MN that arise following ionizing radiation (IR), ruptured MN generated from acute transcription stressors DRB or siSRSF1 are refractory to cGAS localization. Recruitment of cGAS to MN is blocked by inhibiting the histone methyltransferase DOT1L prior to IR exposure, demonstrating that cGAS recruitment to MN is dictated by nuclear chromatin organization at the time of DNA damage. Loss of cGAS+ MN, caused either by acute transcription stressors or by preventing DOT1L-deposited histone methylation, corresponded to significantly decreased cGAS-dependent inflammatory signalling. These results implicate nuclear chromatin organization in micronuclear composition and activity, influencing the ability of damage-induced MN to retain cytosolic proteins upon rupture.

Inside-out Signalling From Aminopeptidase N (CD13) To Complement Receptor 3 (CR3, CD11b/CD18)

Upon ligand engagement, certain receptors can activate an integrin through a mechanism called inside-out signalling. This phenomenon prepares the cell for the next steps of the process it will perform. CR3 (Complement receptor 3), the most abundant β2 integrin in monocytes and macrophages, and CD13 (aminopeptidase N) are two immune receptors with overlapping activities: adhesion, phagocytosis of opsonized particles, and respiratory burst induction. They can be found together in functional signalling microdomains, or lipid rafts, on the surface of human leukocytes. Thus, given their common functions, shared physical location and the fact that some phagocytic and adhesion receptors activate a selection of integrins, we hypothesized that CD13 could activate CR3 through an inside-out signalling mechanism. To test this hypothesis, we first ascertained the activation of CR3 after CD13 crosslinking in human monocyte-derived macrophages. We used an integrated analysis of bioinformatics and experimental data to suggest two possible signalling cascades that could explain the phenomenon. Finally, we show that the non-receptor tyrosine kinase Syk is a key attenuator of this pathway. Our results demonstrated that, even in the absence of canonical signalling motifs, and despite having a noticeably short cytoplasmic tail (7-10 amino acids), CD13 was capable of triggering an inside-out signalling cascade, adding a new function to those already known for this moonlighting protein.

Neurobiology of phenotypic plasticity in the light of climate change

Phenotypic plasticity describes the ability of an organism with a given genotype to respond to changing environmental conditions through the adaptation of the phenotype. Phenotypic plasticity is a widespread means of adaptation, allowing organisms to optimize fitness levels in changing environments. A core prerequisite for adaptive predictive plasticity is the existence of reliable cues, i.e. accurate environmental information about future selection on the expressed plastic phenotype. Furthermore, organisms need the capacity to detect and interpret such cues, relying on specific sensory signalling and neuronal cascades. Subsequent neurohormonal changes lead to the transformation of phenotype A into phenotype B. Each of these activities is critical for survival. Consequently, anything that could impair an animal’s ability to perceive important chemical information could have significant ecological ramifications. Climate change and other human stressors can act on individual or all of the components of this signalling cascade. In consequence, organisms could lose their adaptive potential, or in the worst case, even become maladapted. Therefore, it is key to understand the sensory systems, the neurobiology and the physiological adaptations that mediate organisms’ interactions with their environment. It is, thus, pivotal to predict the ecosystem-wide effects of global human forcing. This review summarizes current insights on how climate change affects phenotypic plasticity, focussing on how associated stressors change the signalling agents, the sensory systems, receptor responses and neuronal signalling cascades, thereby, impairing phenotypic adaptations.

Modulation of TCR signalling components occurs prior to positive selection and lineage commitment in iNKT cells

AbstractiNKT cells play a critical role in controlling the strength and character of adaptive and innate immune responses. Their unique functional characteristics are induced by a transcriptional program initiated by positive selection mediated by CD1d expressed by CD4+CD8+ (double positive, DP) thymocytes. Here, using a novel Vα14 TCR transgenic strain bearing greatly expanded numbers of CD24hiCD44loNKT cells, we examined transcriptional events in four immature thymic iNKT cell subsets. A transcriptional regulatory network approach identified transcriptional changes in proximal components of the TCR signalling cascade in DP NKT cells. Subsequently, positive and negative selection, and lineage commitment, occurred at the transition from DP NKT to CD4 NKT. Thus, this study introduces previously unrecognised steps in early NKT cell development, and separates the events associated with modulation of the T cell signalling cascade prior to changes associated with positive selection and lineage commitment.

The Relative Binding Position of Nck and Grb2 Adaptors Dramatically Impacts Actin-Based Motility of Vaccinia Virus

ABSTRACTPhosphotyrosine (pTyr) motifs in unstructured polypeptides orchestrate important cellular processes by engaging SH2-containing adaptors to nucleate complex signalling networks. The concept of phase separation has recently changed our appreciation of such multivalent networks, however, the role of pTyr motif positioning in their function remains to be explored. We have now explored this parameter in the assembly and operation of the signalling cascade driving actin-based motility and spread of Vaccinia virus. This network involves two pTyr motifs in the viral protein A36 that recruit the adaptors Nck and Grb2 upstream of N-WASP and Arp2/3-mediated actin polymerization. We generated synthetic networks on Vaccinia by manipulating pTyr motifs in A36 and the unrelated p14 from Orthoreovirus. In contrast to predictions, we find that only specific spatial arrangements of Grb2 and Nck binding sites result in robust N-WASP recruitment, Arp2/3 driven actin polymerization and viral spread. Our results suggest that the relative position of pTyr adaptor binding sites is optimised for signal output. This finding may explain why the relative positions of pTyr motifs are usually conserved in proteins from widely different species. It also has important implications for regulation of physiological networks, including those that undergo phase transitions.

Sublytic membrane attack complex drives glycolysis and mitochondrial dysfunction with inflammatory consequences in human monocyte-derived macrophages

The complement system is an ancient and critical element of the innate immune system. The terminal step in the complement pathway, the membrane attack complex (MAC), has previously been linked to inflammasome activation and is linked to the pathogenesis of multiple diseases including rheumatoid arthritis and neurodegeneration. Using both metabolomic and proteomic approaches, here we show that in human monocyte-derived macrophages a sublytic concentration of MAC mediates a previously uncharacterised metabolic shift, mitochondrial dysfunction and upregulation of glycolysis-promoting genes. This skewing of metabolism coupled with mitochondrial dysfunction drives ROS-mediated NLRP3 inflammasome activation and subsequent gasdermin D activation and pro-inflammatory cytokine production and release. Together, these data elucidate a novel immunometabolic signalling cascade in MAC-stimulated human macrophages, the consequences of which have implications in considering much needed novel therapeutic options for diseases linked to aberrant complement activation

Distinct, opposing functions for CFIm59 and CFIm68 in mRNA alternative polyadenylation of Pten and in the PI3K/Akt signalling cascade

ABSTRACTThe precise maintenance of PTEN dosage is crucial for tumor suppression across a wide variety of cancers. Post-transcriptional regulation of Pten heavily relies on regulatory elements encoded by its 3’UTR. We previously reported the important diversity of 3’UTR isoforms of Pten mRNAs produced through alternative polyadenylation (APA). Here, we reveal the direct regulation of Pten APA by the mammalian cleavage factor I (CFIm) complex, which in turn contributes to PTEN protein dosage. CFIm consists of the UGUA-binding CFIm25 and APA regulatory subunits CFIm59 or CFIm68. Deep sequencing analyses of perturbed (KO and KD) cell lines uncovered the differential regulation of Pten APA by CFIm59 and CFIm68 and further revealed that their divergent functions have widespread impact for APA in transcriptomes. Differentially regulated genes include numerous factors within the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signalling pathway that PTEN counter-regulates. We further reveal a stratification of APA dysregulation among a subset of PTEN-driven cancers, with recurrent alterations among PI3K/Akt pathway genes regulated by CFIm. Our results refine the transcriptome selectivity of the CFIm complex in APA regulation, and the breadth of its impact in PTEN-driven cancers.

SUMOylation of Dorsal attenuates Toll/NF-κB signalling

In Drosophila, Toll/NF-κB signalling plays key roles in both animal development and in host defence. The activation, intensity and kinetics of Toll signalling is regulated by post-translational modifications such as phosphorylation, SUMOylation or ubiquitination that target multiple proteins in the Toll/NF-κB cascade.Here, we have generated a CRISPR-Cas9 edited Dorsal (DL) variant that is SUMO conjugation resistant (SCR). Intriguingly, embryos laid by dlSCR mothers overcome dl haploinsufficiency and complete the developmental program. This ability appears to be a result of higher transcriptional activation by DLSCR. In contrast, SUMOylation dampens DL transcriptional activation, ultimately conferring robustness to the dorso-ventral program. In the larval immune response, dlSCR animals show increase in crystal cell numbers, stronger activation of humoral defence genes, high cactus levels and cytoplasmic stabilization of DL:Cactus complexes. A mathematical model that evaluates the contribution of the small fraction of SUMOylated DL (<5%) suggests that it acts to block transcriptional activation, driven primarily by DL that is not SUMO conjugated.Our findings define SUMO conjugation as an important regulator of the Toll signalling cascade, in both development and in host defense. Our results broadly indicate that SUMO attenuates DL at the level of transcriptional activation. Further, we hypothesize that SUMO conjugation of DL may be part of a Ubc9 dependant feedback circuit that restrains Toll/NF-κB signalling.

Transmembrane signalling by a bionic receptor: biological input and output, chemical mechanism of signal transduction

Signal transduction through sealed biological membranes is among the most important evolutionary achievements. Herein, we focus on the development of artificial signal transduction mechanisms and engineer a bionic receptor with capacity of transduction of biological signals across biological membranes using tools of chemistry. The bionic receptor described in this work exhibits similarity with the natural counterpart in the most essential characteristics: in having an exofacial ligand for signal capture, in being membrane anchored, and in featuring a releasable secondary messenger molecule, which performs enzyme activation in the endo volume. The main difference with the natural receptors is that signal transduction across the lipid bilayer was performed using the tools of organic chemistry, namely a self-immolative linker. The highest novelty of our work is that the artificial signalling cascade designed herein achieved transmembrane activation of enzymatic activity, as is the hallmark of activity by natural signalling receptors.

The natural history of defences

Hosts can avoid infections by behavioural changes and by body walls. After infection, hosts can change their behaviours to reduce the effects of parasitism. Immune defences have different arms (humoral or cellular), and organization (innate, adaptive). Innate immunity consists of a collection of different systems that are evolutionarily very old. Adaptive immunity, based on expansion of specific lymphocytes, evolved in the higher vertebrates. Immune defences are regulated tightly and based on receptors that can recognize parasites (or their activity). This triggers a complex signalling cascade that results in the production of further signalling compounds and effectors. Important protein families, e.g. the immunoglobulins, form the molecular backbone. A key to efficient defences is the diversification of receptors, such as the highly evolved somatic diversification processes of advanced adaptive immunity. The microbiota adds to defences in many ways. Immune memory and priming occur throughout the tree of life.