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International audience; SARS-CoV-2 infection results in impaired interferon response in patients with severe COVID-19. However, how SARS-CoV-2 interferes with host immune responses is incompletely understood. Here, we sequence small RNAs from SARS-CoV-2-infected human cells and identify a microRNA (miRNA) derived from a recently evolved region of the viral genome. We show that the virus-derived miRNA produces two miRNA isoforms in infected cells by the enzyme Dicer, which are loaded into Argonaute proteins. Moreover, the predominant miRNA isoform targets the 3'UTR of interferon-stimulated genes and represses their expression in a miRNA-like fashion. Finally, the two viral miRNA isoforms were detected in nasopharyngeal swabs from COVID-19 patients. We propose that SARS-CoV-2 can potentially employ a virus-derived miRNA to hijack the host miRNA machinery, which could help to evade the interferon-mediated immune response.

International audience; Posttranscriptional regulation of gene expression is central to the development and replication of the malaria parasite, Plasmodium falciparum , within its human host. The timely coordination of RNA maturation, homeostasis, and protein synthesis relies on the recruitment of specific RNA-binding proteins to their cognate target mRNAs. One possible mediator of such mRNA-protein interactions is the N 6 -methylation of adenosines (m 6 A), a prevalent mRNA modification of parasite mRNA transcripts. Here, we used RNA protein pulldowns, RNA modification mass spectrometry, and quantitative proteomics to identify two P. falciparum YTH domain proteins (PfYTH.1 and PfYTH.2) as m 6 A-binding proteins during parasite blood-stage development. Interaction proteomics revealed that PfYTH.2 associates with the translation machinery, including multiple subunits of the eukaryotic initiation factor 3 (eIF3) and poly(A)-binding proteins. Furthermore, knock sideways of PfYTH.2 coupled with ribosome profiling showed that this m 6 A reader is essential for parasite survival and is a repressor of mRNA translation. Together, these data reveal an important missing link in the m 6 A-mediated mechanism controlling mRNA translation in a unicellular eukaryotic pathogen.IMPORTANCE Infection with the unicellular eukaryotic pathogen Plasmodium falciparum causes malaria, a mosquito-borne disease affecting more than 200 million and killing 400,000 people each year. Underlying the asexual replication within human red blood cells is a tight regulatory network of gene expression and protein synthesis. A widespread mechanism of posttranscriptional gene regulation is the chemical modification of adenosines (m 6 A), through which the fate of individual mRNA transcripts can be changed. Here, we report on the protein machinery that “reads” this modification and “translates” it into a functional outcome. We provide mechanistic insight into one m 6 A reader protein and show that it interacts with the translational machinery and acts as a repressor of mRNA translation. This m 6 A-mediated phenotype has not been described in other eukaryotes as yet, and the functional characterization of the m 6 A interactome will ultimately open new avenues to combat the disease.

Posttranscriptional regulation of gene expression is central to the development and replication of the malaria parasite, Plasmodium falciparum, within its human host. The timely coordination of RNA maturation, homeostasis, and protein synthesis relies on the recruitment of specific RNA-binding proteins to their cognate target mRNAs. One possible mediator of such mRNA-protein interactions is the N6-methylation of adenosines (m6A), a prevalent mRNA modification of parasite mRNA transcripts. Here, we used RNA protein pulldowns, RNA modification mass spectrometry, and quantitative proteomics to identify two P. falciparum YTH domain proteins (PfYTH.1 and PfYTH.2) as m6A-binding proteins during parasite blood-stage development. Interaction proteomics revealed that PfYTH.2 associates with the translation machinery, including multiple subunits of the eukaryotic initiation factor 3 (eIF3) and poly(A)-binding proteins. Furthermore, knock sideways of PfYTH.2 coupled with ribosome profiling showed that this m6A reader is essential for parasite survival and is a repressor of mRNA translation. Together, these data reveal an important missing link in the m6A-mediated mechanism controlling mRNA translation in a unicellular eukaryotic pathogen.IMPORTANCE Infection with the unicellular eukaryotic pathogen Plasmodium falciparum causes malaria, a mosquito-borne disease affecting more than 200 million and killing 400,000 people each year. Underlying the asexual replication within human red blood cells is a tight regulatory network of gene expression and protein synthesis. A widespread mechanism of posttranscriptional gene regulation is the chemical modification of adenosines (m6A), through which the fate of individual mRNA transcripts can be changed. Here, we report on the protein machinery that "reads" this modification and "translates" it into a functional outcome. We provide mechanistic insight into one m6A reader protein and show that it interacts with the translational machinery and acts as a repressor of mRNA translation. This m6A-mediated phenotype has not been described in other eukaryotes as yet, and the functional characterization of the m6A interactome will ultimately open new avenues to combat the disease. Ministry of Education (MOE) National Research Foundation (NRF) Singapore-MIT Alliance for Research and Technology (SMART) Published version This work was supported by a European Research Council Advanced Grant (PlasmoSilencing 670301) and the French Parasitology consortium ParaFrap (ANR-11- LABX0024) awarded to A. Scherf. S.B. is supported by an EMBO Advanced Long-Term Fellowship (aALTF 632-2018). P.R.P., P.C.D., J.L., and A. Sinha were supported by the National Research Foundation Singapore under its Singapore-MIT Alliance for Research and Technology (SMART) Centre, Antimicrobial Resistance IRG and Singapore Ministry of Education Academic Research Fund Tier 2 (MOE2018-T2-2-131). A. Sinha and J.L. acknowledge support from the Singapore-MIT Alliance (SMA) Graduate Fellowship. Proteomics work was performed in part in the Center for Environmental Health Sciences BioCore, which is supported by Center grant P30-ES002109 from the National Institute of Environmental Health Sciences. We also acknowledge Peiying Ho and Tan Tse Mien for providing support at SMART laboratories in Singapore. G.C. and M.S. were funded by a European Research Council (ERC) Starting Grant (ERC-StG-679243). Work on this project in the laboratory of S.A.R. is funded by a grant from the Australian National Health and Medical Research Council (1165354) and by a grant from the Australian Research Council (DP160100389).

Argonaute proteins and their interacting small RNAs play a key role in regulating complementary mRNA targets during animal development. Here, we investigate a novel and essential function of the catalytically active Argonaute protein CSR-1 in maternal mRNA degradation during early embryogenesis in Caenorhabditis elegans . We show that CSR-1 interacts with endogenous small RNAs antisense to hundreds of cleared maternal mRNAs in embryos, and preferentially cleaves mRNAs no longer engaged in translation. The depletion of CSR-1 during maternal to zygotic transition leads to embryonic lethality in a catalytic-dependent manner and impairs the degradations of its embryonic mRNA targets. Given the conservation of Argonaute catalytic activity, we propose that a similar mechanism operate to clear maternal mRNAs during maternal to zygotic transition across species.

HSP90 chaperones are essential regulators of cellular function, as they ensure the appropriate conformation of multiple key client proteins. Four HSP90 isoforms were identified in the protozoan parasite Theileria annulata. Partial characterization was undertaken for three and localization confirmed for cytoplasmic (TA12105), endoplasmic reticulum (TA06470), and apicoplast (TA10720) forms. ATPase activity and binding to the HSP90 inhibitor geldanamycin were demonstrated for recombinant TA12105, and all three native forms could be isolated to varying extents by binding to geldanamycin beads. Because it is essential, HSP90 is considered a potential therapeutic drug target. Resistance to the only specific Theileriacidal drug is increasing, and one challenge for design of drugs that target the parasite is to limit the effect on the host. An in vitro cell culture system that allows comparison between uninfected bovine cells and the T. annulata‐infected counterpart was utilized to test the effects of geldanamycin and the derivative 17‐AAG. T. annulata‐infected cells had greater tolerance to geldanamycin than uninfected cells yet exhibited significantly more sensitivity to 17‐AAG. These findings suggest that parasite HSP90 isoform(s) can alter the drug sensitivity of infected host cells and that members of the Theileria HSP90 family are potential targets worthy of further investigation.

International audience; Eukaryotic genomes harbor invading transposable elements that are silenced by PIWI-interacting RNAs (piRNAs) to maintain genome integrity in animal germ cells. However, whether piRNAs also regulate endogenous gene expression programs remains unclear. Here, we show that C. elegans piRNAs trigger the transcriptional silencing of hundreds of spermatogenic genes during spermatogenesis, promoting sperm differentiation and function. This silencing signal requires piRNA-dependent small RNA biogenesis and loading into downstream nuclear effectors, which correlates with the dynamic reorganization of two distinct perinuclear biomolecular condensates present in germ cells. In addition, the silencing capacity of piRNAs is temporally counteracted by the Argonaute CSR-1, which targets and licenses spermatogenic gene transcription. The spatial and temporal overlap between these opposing small RNA pathways contributes to setting up the timing of the spermatogenic differentiation program. Thus, our work identifies a prominent role for piRNAs as direct regulators of endogenous transcriptional programs during germline development and gamete differentiation.