Death domain-associated protein 6 (Daxx) is a multifunctional, ubiquitously expressed and highly conserved chaperone protein involved in numerous cellular processes, including apoptosis, transcriptional repression, and carcinogenesis. In 2015, we identified Daxx as an antiretroviral factor that interfered with HIV-1 replication by inhibiting the reverse transcription step. In the present study, we sought to unravel the molecular mechanism of Daxx-mediated restriction and, in particular, to identify the protein(s) that Daxx targets in order to achieve its antiviral activity. First, we show that the SUMO-interacting motif (SIM) located at the C-terminus of the protein is strictly required for Daxx to inhibit HIV-1 reverse transcription. By performing a quantitative proteomic screen combined with classical biochemical analyses, we found that Daxx associated with incoming HIV-1 cores through a SIM-dependent interaction with cyclophilin A (CypA) and capsid (CA). Daxx was found to reside within a multiprotein complex associated with viral capsids, also containing TNPO3, TRIM5α, and TRIM34. Given the well-known influence of these cellular factors on the stability of HIV-1 cores, we investigated the effect of Daxx on the cytoplasmic fate of incoming cores and found that Daxx prevented HIV-1 uncoating in a SIM-dependent manner. Altogether, our findings suggest that, by recruiting TNPO3, TRIM5α, and TRIM34 and possibly other proteins onto incoming HIV-1 cores through a SIM-dependent interaction with CA-bound CypA, Daxx increases their stability, thus preventing uncoating and reverse transcription. Our study uncovers a previously unknown function of Daxx in the early steps of HIV-1 infection and further illustrates how reverse transcription and uncoating are two tightly interdependent processes.

Spider venoms represent an original source of novel compounds with therapeutic and agrochemical potential. Whereas most of the research efforts have focused on large mygalomorph spiders, araneomorph spiders are equally promising but require more sensitive and sophisticated approaches given their limited size and reduced venom yield. Belonging to the latter group, the genus Lycosa (“wolf spiders”) contains many species widely distributed throughout the world. These spiders are ambush predators that do not build webs but instead rely strongly on their venom for prey capture. Lycosa tarantula is one of the largest species of wolf spider, but its venom composition is unknown. Using a combination of RNA sequencing of the venom glands and venom proteomics, we provide the first overview of the peptides and proteins produced by this iconic Mediterranean spider. Beside the typical small disulfide rich neurotoxins, several families of proteins were also identified, including cysteine-rich secretory proteins (CRISP) and Hyaluronidases. Proteomic analysis of the electrically stimulated venom validated 30 of these transcriptomic sequences, including nine putative neurotoxins and eight venom proteins. Interestingly, LC-MS venom profiles of manual versus electric stimulation, as well as female versus male, showed some marked differences in mass distribution. Finally, we also present some preliminary data on the biological activity of L. tarantula crude venom.

The success of oral infection by viruses depends on their capacity to overcome the gut epithelial barrier of their host to crossing over apical, mucous extracellular matrices. As orally transmitted viruses, densoviruses, are also challenged by the complexity of the insect gut barriers, more specifically by the chitinous peritrophic matrix, that lines and protects the midgut epithelium; how capsids stick to and cross these barriers to reach their final cell destination where replication goes has been poorly studied in insects. Here, we analyzed the early interaction of the Junonia coenia densovirus (JcDV) with the midgut barriers of caterpillars from the pest Spodoptera frugiperda. Using combination of imaging, biochemical, proteomic and transcriptomic analyses, we examined in vitro, ex vivo and in vivo the early interaction of the capsids with the peritrophic matrix and the consequence of early oral infection on the overall gut function. We show that the JcDV particle rapidly adheres to the peritrophic matrix through interaction with different glycans including chitin and glycoproteins, and that these interactions are necessary for oral infection. Proteomic analyses of JcDV binding proteins of the peritrophic matrix revealed mucins and non-mucins proteins including enzymes already known to act as receptors for several insect pathogens. In addition, we show that JcDV early infection results in an arrest of N-Acetylglucosamine secretion and a disruption in the integrity of the peritrophic matrix, which may help viral particles to pass through. Finally, JcDV early infection induces changes in midgut genes expression favoring an increased metabolism including an increased translational activity. These dysregulations probably participate to the overall dysfunction of the gut barrier in the early steps of viral pathogenesis. A better understanding of early steps of densovirus infection process is crucial to build biocontrol strategies against major insect pests.

The frequency of synonymous codons in protein coding genes is non-random and varies both between species and between genes within species. Whether this codon usage bias (CUBias) reflects underlying neutral mutational processes or is instead shaped by selection remains an open debate, especially regarding the role of selection for enhanced protein production. Variation in CUBias of a gene (be it natural synonymous mutations or biotechnological synonymous recoding) can have an enormous impact on its expression by diverse cis -acting mechanisms. But expression of genes with extreme CUBias can also lead to strong phenotypic effects by altering the overall intracellular translation homeostasis via competition for ribosomal machinery or tRNA depletion. In this study, we expressed at high levels in human cells six different synonymous versions of a gene and used matched transcriptomic and proteomic data to evaluate the impact of CUBias of the heterologous gene on the translation of cellular transcripts. Our experimental design focused specifically on differences during translation elongation. Response to expression of the different synonymous sequences was assessed by various approaches, ranging from analyses performed on a per-gene basis to more integrated approaches of the cell as a whole. We observe that the transcriptome displayed substantial changes as a result of heterologous gene expression by triggering an intense antiviral and inflammatory response, but that changes in the proteomes were very modest. Most importantly we notice that changes in translation efficiency of cellular transcripts were not associated with the direction of the CUBias of the heterologous sequences, thereby providing only limited support for trans -acting effects of synonymous changes. We interpret that, in human cells in culture, changes in CUBias can lead to important cis -acting effects in gene expression, but that cellular homeostasis can buffer the phenotypic impact of overexpression of heterologous genes with extreme CUBias.

Summary: Organoids are powerful models of brain development and function. Mouse brain organoids reproduce the main steps of neurodevelopment in vivo and differentiate more rapidly than human organoids, making them scalable for screening applications. However, mouse brain organoids remain poorly defined. Here, we showed that the transcriptome of mouse brain organoids developed for three weeks is close to that of a neonatal mouse brain. Strikingly, organoids reproduced the majority of alternative splicing and polyadenylation site events that define a neural identity in vivo . Proteomics revealed that most changes in mRNA expression were translated into proteins. Mouse brain organoids differentiated for only three weeks already harboured a complex network of synaptic proteins, including the ionotropic and metabotropic receptors for the neurotransmitters glutamate and GABA. To conclude, our study provides compelling evidence that mouse brain organoids are a rapidly maturing and relevant model of mammalian brain development and function in a neonatal-like environment.

All the members of the phylum Cnidaria are characterized by the production of venom in specialized structures, the nematocysts. Venom of jellyfish (Medusozoa) and sea anemones (Anthozoa) has been investigated since the 1970s, revealing a remarkable molecular diversity. Specifically, sea anemones harbour a rich repertoire of neurotoxic peptides, some of which have been developed in drug leads. However, venoms of the vast majority of Anthozoa species remain uncharacterized, particularly in the class Octocorallia. To fill this gap, we applied a proteo-transcriptomic approach to investigate venom composition in Eunicella singularis , a gorgonian species common in Mediterranean hard-bottom benthic communities. Our results highlighted the peculiarities of the venom of E. singularis with respect to sea anemones, which is reflected in the presence of several toxins with novel folds, worthy of functional characterization. A comparative genomic survey across the octocoral radiation allowed us to generalize these findings and provided insights into the evolutionary history, molecular diversification patterns and putative adaptive roles of venom toxins. A comparison of whole-body and nematocyst proteomes revealed the presence of different cytolytic toxins inside and outside the nematocysts. Two instances of differential maturation patterns of toxin precursors were also identified, highlighting the intricate regulatory pathways underlying toxin expression.