A liver immune rheostat regulates CD8 T cell immunity in chronic HBV infection.

Chronic hepatitis B virus (HBV) infection affects 300 million patients worldwide1,2, in whom virus-specific CD8 T cells by still ill-defined mechanisms lose their function and cannot eliminate HBV-infected hepatocytes3-7. Here we demonstrate that a liver immune rheostat renders virus-specific CD8 T cells refractory to activation and leads to their loss of effector functions. In preclinical models of persistent infection with hepatotropic viruses such as HBV, dysfunctional virus-specific CXCR6+ CD8 T cells accumulated in the liver and, as a characteristic hallmark, showed enhanced transcriptional activity of cAMP-responsive element modulator (CREM) distinct from T cell exhaustion. In patients with chronic hepatitis B, circulating and intrahepatic HBV-specific CXCR6+ CD8 T cells with enhanced CREM expression and transcriptional activity were detected at a frequency of 12-22% of HBV-specific CD8 T cells. Knocking out the inhibitory CREM/ICER isoform in T cells, however, failed to rescue T cell immunity. This indicates that CREM activity was a consequence, rather than the cause, of loss in T cell function, further supported by the observation of enhanced phosphorylation of protein kinase A (PKA) which is upstream of CREM. Indeed, we found that enhanced cAMP-PKA-signalling from increased T cell adenylyl cyclase activity augmented CREM activity and curbed T cell activation and effector function in persistent hepatic infection. Mechanistically, CD8 T cells recognizing their antigen on hepatocytes established close and extensive contact with liver sinusoidal endothelial cells, thereby enhancing adenylyl cyclase-cAMP-PKA signalling in T cells. In these hepatic CD8 T cells, which recognize their antigen on hepatocytes, phosphorylation of key signalling kinases of the T cell receptor signalling pathway was impaired, which rendered them refractory to activation. Thus, close contact with liver sinusoidal endothelial cells curbs the activation and effector function of HBV-specific CD8 T cells that target hepatocytes expressing viral antigens by means of the adenylyl cyclase-cAMP-PKA axis in an immune rheostat-like fashion.

Genome-scale pan-cancer interrogation of lncRNA dependencies using CasRx.

Although long noncoding RNAs (lncRNAs) dominate the transcriptome, their functions are largely unexplored. The extensive overlap of lncRNAs with coding and regulatory sequences restricts their systematic interrogation by DNA-directed perturbation. Here we developed genome-scale lncRNA transcriptome screening using Cas13d/CasRx. We show that RNA targeting overcomes limitations inherent to other screening methods, thereby considerably expanding the explorable space of the lncRNAome. By evolving the screening system toward pan-cancer applicability, it supports molecular and phenotypic data integration to contextualize screening hits or infer lncRNA function. We thereby addressed challenges posed by the enormous transcriptome size and tissue specificity through a size-reduced multiplexed gRNA library termed Albarossa, targeting 24,171 lncRNA genes. Its rational design incorporates target prioritization based on expression, evolutionary conservation and tissue specificity, thereby reconciling high discovery power and pan-cancer representation with scalable experimental throughput. Applied across entities, the screening platform identified numerous context-specific and common essential lncRNAs. Our work sets the stage for systematic exploration of lncRNA biology in health and disease.

Mutual regulation of CD4+ T cells and intravascular fibrin in infections.

Innate myeloid cells especially neutrophils and their extracellular traps are known to promote intravascular coagulation and thrombosis formation in infections and various other conditions. Innate myeloid cell-dependent fibrin formation can support systemic immunity while its dysregulation enhances the severity of infectious diseases. Less is known about the immune mechanisms preventing dysregulation of fibrin homeostasis in infection. During experimental systemic infections local fibrin deposits in the liver microcirculation cause rapid arrest of CD4+ T cells. Arrested T-helper cells mostly represent Th17 cells that partially originate from the small intestine. Intravascular fibrin deposits activate mouse and human CD4+ T cells which can be mediated by direct fibrin-CD4+ T-cell interactions. Activated CD4+ T cells suppress fibrin deposition and microvascular thrombosis by directly counteracting coagulation activation by neutrophils and classical monocytes. T-cell activation, which is initially triggered by IL-12p40- and MHC-II-dependent mechanisms, enhances intravascular fibrinolysis via LFA-1. Moreover, CD4+ T cells disfavor the association of the thrombin-activatable fibrinolysis inhibitor (TAFI) with fibrin whereby fibrin deposition is increased by TAFI in the absence but not in the presence of T cells. In human infections thrombosis development is inversely related to microvascular levels of CD4+ T cells. Thus, fibrin promotes LFA-1-dependent T-helper cell activation in infections which drives a negative feedback cycle that rapidly restricts intravascular fibrin and thrombosis development.