EMBO Reports最新文献

Genome transcription and replication of influenza A virus (FluA), catalyzed by viral RNA polymerase (FluAPol), are delicately controlled across the virus life cycle. A switch from transcription to replication occurring at later stage of an infection is critical for progeny virion production and viral non-structural protein NS2 has been implicated in regulating the switch. However, the underlying regulatory mechanisms and the structure of NS2 remained elusive for years. Here, we determine the cryo-EM structure of the FluAPol-NS2 complex at ~3.0 Å resolution. Surprisingly, three domain-swapped NS2 dimers arrange three symmetrical FluPol dimers into a highly ordered barrel-like hexamer. Further structural and functional analyses demonstrate that NS2 binding not only hampers the interaction between FluAPol and the Pol II CTD because of steric conflicts, but also impairs FluAPol transcriptase activity by stalling it in the replicase conformation. Moreover, this is the first visualization of the full-length NS2 structure. Our findings uncover key molecular mechanisms of the FluA transcription-replication switch and have implications for the development of antivirals.

Phenotypic plasticity is a cause of glioblastoma therapy failure. We previously showed that suppressing the oligodendrocyte-lineage regulator SOX10 promotes glioblastoma progression. Here, we analyze SOX10-mediated phenotypic plasticity and exploit it for glioblastoma therapy design. We show that low SOX10 expression is linked to neural stem-cell (NSC)-like glioblastoma cell states and is a consequence of temozolomide treatment in animal and cell line models. Single-cell transcriptome profiling of Sox10-KD tumors indicates that Sox10 suppression is sufficient to induce tumor progression to an aggressive NSC/developmental-like phenotype, including a quiescent NSC-like cell population. The quiescent NSC state is induced by temozolomide and Sox10-KD and reduced by Notch pathway inhibition in cell line models. Combination treatment using Notch and HDAC/PI3K inhibitors extends the survival of mice carrying Sox10-KD tumors, validating our experimental therapy approach. In summary, SOX10 suppression mediates glioblastoma progression through NSC/developmental cell-state transition, including the induction of a targetable quiescent NSC state. This work provides a rationale for the design of tumor therapies based on single-cell phenotypic plasticity analysis.

Epithelial-immune cell communication is pivotal to control microbial infections. We show that glycosylphosphatidylinositol-linked aspartyl proteases (Yapsins) of the human opportunistic pathogenic yeast Candida glabrata (Cg) thwart epithelial cell (EC)-neutrophil signalling by targeting the EC protein, Arpc1B (actin nucleator Arp2/3 complex subunit), which leads to actin disassembly and impeded IL-8 secretion by ECs. Further, the diminished IL-8 secretion inhibits neutrophil migration, and protects Cg from the neutrophil-mediated killing. CgYapsin-dependent Arpc1B degradation requires Arginine-142 in Arpc1B, and leads to reduced Arpc1B-p38 MAPK interaction and downregulated p38 signalling. Consistently, Arpc1B or p38 deletion promotes survival of the Cg aspartyl protease-deficient mutant in ECs. Importantly, kidneys of the protease-deficient mutant-infected mice display elevated immune cell infiltration and cytokine secretion, implicating CgYapsins in immune response suppression in vivo. Besides delineating Cg-EC interplay, our results uncover a novel target, Arpc1B, that pathogens attack to constrain the host signalling networks, and link Arpc1B mechanistically with p38 activation.

TLR8 senses single-stranded RNA (ssRNA) fragments, processed via cleavage by ribonuclease (RNase) T2 and RNase A family members. Processing by these RNases releases uridines and purine-terminated residues resulting in TLR8 activation. Monocytes show high expression of RNase 6, yet this RNase has not been analyzed for its physiological contribution to the recognition of bacterial RNA by TLR8. Here, we show a role for RNase 6 in TLR8 activation. BLaER1 cells, transdifferentiated into monocyte-like cells, as well as primary monocytes deficient for RNASE6 show a dampened TLR8-dependent response upon stimulation with isolated bacterial RNA (bRNA) and also upon infection with live bacteria. Pretreatment of bacterial RNA with recombinant RNase 6 generates fragments that induce TLR8 stimulation in RNase 6 knockout cells. 2'O-RNA methyl modification, when introduced at the first uridine in the UA dinucleotide, impairs processing by RNase 6 and dampens TLR8 stimulation. In summary, our data show that RNase 6 processes bacterial RNA and generates uridine-terminated breakdown products that activate TLR8.

Tumor-associated macrophages (TAMs) are prime therapeutic targets due to their pro-tumorigenic functions, but varying efficacy of macrophage-targeting therapies highlights our incomplete understanding of how macrophages are regulated within the tumor microenvironment (TME). The circadian clock is a key regulator of macrophage function, but how circadian rhythms of macrophages are influenced by the TME remains unknown. Here, we show that conditions associated with the TME such as polarizing stimuli, acidic pH, and lactate can alter circadian rhythms in macrophages. While cyclic AMP (cAMP) has been reported to play a role in macrophage response to acidic pH, our results indicate pH-driven changes in circadian rhythms are not mediated solely by cAMP signaling. Remarkably, circadian disorder of TAMs was revealed by clock correlation distance analysis. Our data suggest that heterogeneity in circadian rhythms within the TAM population level may underlie this circadian disorder. Finally, we report that circadian regulation of macrophages suppresses tumor growth in a murine model of pancreatic cancer. Our work demonstrates a novel mechanism by which the TME influences macrophage biology through modulation of circadian rhythms.

Wingless (Wg)/Wnt signaling plays a critical role in both development and adult tissue homeostasis. In the Drosophila larval wing disc epithelium, the orderly delivery of Wg/Wnt to the apical and basal cell surfaces is essential for wing development. Here, we identified Ehbp1 as the switch that dictates the direction of Wg/Wnt polarized intracellular transport: the Adaptor Protein complex 1 (AP-1) delivers Wg/Wnt to the basolateral cell surface, and its sequestration by Ehbp1 redirects Wg/Wnt for apical delivery. Genetic analyses showed that Ehbp1 specifically regulates the polarized distribution of Wg/Wnt, a process that depends on the dedicated Wg/Wnt cargo receptor Wntless. Mechanistically, Ehbp1 competes with Wntless for AP-1 binding, thereby preventing the unregulated basolateral Wg/Wnt transport. Reducing Ehbp1 expression, or removing the coiled-coil motifs within its bMERB domain, leads to basolateral Wg/Wnt accumulation. Importantly, the regulation of polarized Wnt delivery by EHBP1 is conserved in vertebrates. The generality of this switch mechanism for regulating intracellular transport remains to be determined in future studies.

Dinoflagellates, a class of unicellular eukaryotic phytoplankton, exhibit minimal transcriptional regulation, representing a unique model for exploring gene expression. The biosynthesis, distribution, regulation, and function of mRNA N1-methyladenosine (m¹A) remain controversial due to its limited presence in typical eukaryotic mRNA. This study provides a comprehensive map of m¹A in dinoflagellate mRNA and shows that m¹A, rather than N6-methyladenosine (m⁶A), is the most prevalent internal mRNA modification in various dinoflagellate species, with an asymmetric distribution along mature transcripts. In Amphidinium carterae, we identify 6549 m¹A sites characterized by a non-tRNA T-loop-like sequence motif within the transcripts of 3196 genes, many of which are involved in regulating carbon and nitrogen metabolism. Enriched within 3'UTRs, dinoflagellate mRNA m¹A levels negatively correlate with translation efficiency. Nitrogen depletion further decreases mRNA m¹A levels. Our data suggest that distinctive patterns of m¹A modification might influence the expression of metabolism-related genes through translational control.

Morphogens, locally produced signaling molecules, form a concentration gradient to guide tissue patterning. Tissue patterns emerge as a collaboration between morphogen diffusion and responsive cell behaviors, but the mechanisms through which diffusing morphogens define precise spatial patterns amidst biological fluctuations remain unclear. To investigate how cells respond to diffusing proteins to generate tissue patterns, we develop SYMPLE3D, a 3D culture platform. By engineering gene expression responsive to artificial morphogens, we observe that coupling morphogen signals with cadherin-based adhesion is sufficient to convert a morphogen gradient into distinct tissue domains. Morphogen-induced cadherins gather activated cells into a single domain, removing ectopically activated cells. In addition, we reveal a switch-like induction of cadherin-mediated compaction and cell mixing, homogenizing activated cells within the morphogen gradient to form a uniformly activated domain with a sharp boundary. These findings highlight the cooperation between morphogen gradients and cell adhesion in robust tissue patterning and introduce a novel method for tissue engineering to develop new tissue domains in organoids.