A hierarchical assembly pathway directs the unique subunit arrangement of TRiC/CCT.

How the essential eukaryotic chaperonin TRiC/CCT assembles from eight distinct subunits into a unique double-ring architecture remains undefined. We show TRiC assembly involves a hierarchical pathway that segregates subunits with distinct functional properties until holocomplex (HC) completion. A stable, likely early intermediate arises from small oligomers containing CCT2, CCT4, CCT5, and CCT7, contiguous subunits that constitute the negatively charged hemisphere of the TRiC chamber, which has weak affinity for unfolded actin. The remaining subunits CCT8, CCT1, CCT3, and CCT6, which comprise the positively charged chamber hemisphere that binds unfolded actin more strongly, join the ring individually. Unincorporated late-assembling subunits are highly labile in cells, which prevents their accumulation and premature substrate binding. Recapitulation of assembly in a recombinant system demonstrates that the subunits in each hemisphere readily form stable, noncanonical TRiC-like HCs with aberrant functional properties. Thus, regulation of TRiC assembly along a biochemical axis disfavors the formation of stable alternative chaperonin complexes.

Cotranslational prolyl hydroxylation is essential for flavivirus biogenesis.

Viral pathogens are an ongoing threat to public health worldwide. Analysing their dependence on host biosynthetic pathways could lead to effective antiviral therapies1. Here we integrate proteomic analyses of polysomes with functional genomics and pharmacological interventions to define how enteroviruses and flaviviruses remodel host polysomes to synthesize viral proteins and disable host protein production. We find that infection with polio, dengue or Zika virus markedly modifies polysome composition, without major changes to core ribosome stoichiometry. These viruses use different strategies to evict a common set of translation initiation and RNA surveillance factors from polysomes while recruiting host machineries that are specifically required for viral biogenesis. Targeting these specialized viral polysomes could provide a new approach for antiviral interventions. For example, we find that both Zika and dengue use the collagen proline hydroxylation machinery to mediate cotranslational modification of conserved proline residues in the viral polyprotein. Genetic or pharmacological inhibition of proline hydroxylation impairs nascent viral polyprotein folding and induces its aggregation and degradation. Notably, such interventions prevent viral polysome remodelling and lower virus production. Our findings delineate the modular nature of polysome specialization at the virus-host interface and establish a powerful strategy to identify targets for selective antiviral interventions.

The cumulative incidence of cisplatin-induced hearing loss in young children is higher and develops at an early stage during therapy compared with older children based on 2052 audiological assessments.

BACKGROUND: Ototoxicity is a common adverse event of cisplatin treatment. The authors investigated the development of cisplatin-induced hearing loss (CIHL) over time in children with cancer by age and examined the influence of other clinical characteristics on the course of CIHL. METHODS: Data from Canadian patients with childhood cancer were retrospectively reviewed. Hearing loss was graded according to International Society of Pediatric Oncology criteria. The Kaplan-Meier method was applied to estimate the cumulative incidence of CIHL for the total cohort and according to age. Cox regression models were used to explore the effects of independent variables on CIHL development up to 3 years after the start of therapy. RESULTS: In total, 368 patients with 2052 audiological assessments were included. Three years after initiating therapy, the cumulative incidence of CIHL was highest in patients aged ≤5 years (75%; 95% confidence interval [CI], 66%-84%), with a rapid increase observed to 27% (95% CI, 21%-35%) at 3 months and to 61% (95% CI, 53%-69%) at 1 year, compared with patients aged >5 years (48%; 95% CI, 37%-62%; P < .001). The total cumulative dose of cisplatin at 3 months (per 100 mg/m2 increase: hazard ratio [HR], 1.20; 95% CI, 1.01-1.41) vincristine (HR, 2.87; 95% CI, 1.89-4.36) and the total duration of concomitantly administered antibiotics (>30 days: HR, 1.85; 95% CI, 1.17-2.95) further influenced CIHL development over time. CONCLUSIONS: In young children, the cumulative incidence of CIHL is higher compared with that in older children and develops early during therapy. The course of CIHL is further influenced by the total cumulative dose of cisplatin and other ototoxic (co-)medication. These results highlight the need for audiological monitoring at each cisplatin cycle.

Patient-specific genetic factors predict treatment failure in sofosbuvir-treated patients with chronic hepatitis C.

BACKGROUND & AIMS: According to pivotal clinical trials, cure rates for sofosbuvir-based antiviral therapy exceed 96%. Treatment failure is usually assumed to be because of virological resistance-associated substitutions or clinical risk factors, yet the role of patient-specific genetic factors has not been well explored. We determined if patient-specific genetic factors help predict patients likely to fail sofosbuvir treatment in real-world treatment situations. METHODS: We recruited sofosbuvir-treated patients with chronic hepatitis C from five Canadian treatment sites, and performed a case-control pharmacogenomics study assessing both previously published and novel genetic polymorphisms. Specifically studied were variants predicted to impair CES1-dependent production of sofosbuvir's active metabolite, interferon-λ signalling variants expected to impact a patient's immune response to the virus and an HLA variant associated with increased spontaneous and treatment-induced viral clearance. RESULTS: Three hundred and fifty-nine sofosbuvir-treated patients were available for analyses after exclusions, with 34 (9.5%) failing treatment. We identified CES1 variants as novel predictors for treatment failure in European patients (rs115629050 or rs4513095; odds ratio (OR): 5.43; 95% confidence interval (CI): 1.64-18.01; P = .0057), replicated associations with IFNL4 variants predicted to increase interferon-λ signalling (eg rs12979860; OR: 2.25; 95% CI: 1.25-4.06; P = .0071) and discovered a novel association with a coding variant predicted to enhance the activity of IFNL4's receptor (rs2834167 in IL10RB; OR: 1.81; 95% CI: 1.01-3.24; P = .047). CONCLUSIONS: Ultimately, this work demonstrates that patient-specific genetic factors could be used as a tool to identify patients at higher risk of treatment failure and allow for these patients to receive effective therapy sooner.

Chemically induced vesiculation as a platform for studying TMEM16F activity.

Calcium-activated phospholipid scramblase mediates the energy-independent bidirectional translocation of lipids across the bilayer, leading to transient or, in the case of apoptotic scrambling, sustained collapse of membrane asymmetry. Cells lacking TMEM16F-dependent lipid scrambling activity are deficient in generation of extracellular vesicles (EVs) that shed from the plasma membrane in a Ca2+-dependent manner, namely microvesicles. We have adapted chemical induction of giant plasma membrane vesicles (GPMVs), which require both TMEM16F-dependent phospholipid scrambling and calcium influx, as a kinetic assay to investigate the mechanism of TMEM16F activity. Using the GPMV assay, we identify and characterize both inactivating and activating mutants that elucidate the mechanism for TMEM16F activation and facilitate further investigation of TMEM16F-mediated lipid translocation and its role in extracellular vesiculation.

Tau/MAPT disease-associated variant A152T alters tau function and toxicity via impaired retrograde axonal transport.

Mutations in the microtubule-associated protein tau (MAPT) underlie multiple neurodegenerative disorders, yet the pathophysiological mechanisms are unclear. A novel variant in MAPT resulting in an alanine to threonine substitution at position 152 (A152T tau) has recently been described as a significant risk factor for both frontotemporal lobar degeneration and Alzheimer's disease. Here we use complementary computational, biochemical, molecular, genetic and imaging approaches in Caenorhabditis elegans and mouse models to interrogate the effects of the A152T variant on tau function. In silico analysis suggests that a threonine at position 152 of tau confers a new phosphorylation site. This finding is borne out by mass spectrometric survey of A152T tau phosphorylation in C. elegans and mouse. Optical pulse-chase experiments of Dendra2-tau demonstrate that A152T tau and phosphomimetic A152E tau exhibit increased diffusion kinetics and the ability to traverse across the axon initial segment more efficiently than wild-type (WT) tau. A C. elegans model of tauopathy reveals that A152T and A152E tau confer patterns of developmental toxicity distinct from WT tau, likely due to differential effects on retrograde axonal transport. These data support a role for phosphorylation of the variant threonine in A152T tau toxicity and suggest a mechanism involving impaired retrograde axonal transport contributing to human neurodegenerative disease.

Phosphorylation at distinct subcellular locations underlies specificity in mTORC2-mediated activation of SGK1 and Akt.

mTORC2 lies at the intersection of signaling pathways that control metabolism and ion transport through phosphorylation of the AGC-family kinases, the Akt and SGK1 proteins. How mTORC2 targets these functionally distinct downstream effectors in a context-specific manner is not known. Here, we show that the salt- and blood pressure-regulatory hormone, angiotensin II (AngII) stimulates selective mTORC2-dependent phosphorylation of SGK1 (S422) but not Akt (S473 and equivalent sites). Conventional PKC (cPKC), a critical mediator of the angiotensin type I receptor (AT1R, also known as AGTR1) signaling, regulates the subcellular localization of SIN1 (also known as MAPKAP1) and SGK1. Inhibition of cPKC catalytic activity disturbs SIN1 and SGK1 subcellular localization, re-localizing them from the nucleus and a perinuclear compartment to the plasma membrane in advance of hormonal stimulation. Surprisingly, pre-targeting of SIN1 and SGK1 to the plasma membrane prevents SGK1 S422 but not Akt S473 phosphorylation. Additionally, we identify three sites on SIN1 (S128, S315 and S356) that are phosphorylated in response to cPKC activation. Collectively, these data demonstrate that SGK1 activation occurs at a distinct subcellular compartment from that of Akt and suggests a mechanism for the selective activation of these functionally distinct mTORC2 targets through subcellular partitioning of mTORC2 activity.

Mechanisms underlying the synaptic trafficking of the glutamate delta receptor GluD1.

Ionotropic glutamate delta receptors do not bind glutamate and do not generate ionic current, resulting in difficulty in studying the function and trafficking of these receptors. Here, we utilize chimeric constructs, in which the ligand-binding domain of GluD1 is replaced by that of GluK1, to examine its synaptic trafficking and plasticity. GluD1 trafficked to the synapse, but was incapable of expressing long-term potentiation (LTP). The C-terminal domain (CT) of GluD1 has a classic PDZ-binding motif, which is critical for the synaptic trafficking of other glutamate receptors, but we found that its binding to PSD-95 was very weak, and deleting the PDZ-binding motif failed to alter synaptic trafficking. However, deletion of the entire CT abolished synaptic trafficking, but not surface expression. We found that mutation of threonine (T) T923 to an alanine disrupted synaptic trafficking. Therefore, GluD1 receptors have strikingly different trafficking mechanisms compared with AMPARs. These results highlight the diversity of ionotropic glutamate receptor trafficking rules at a single type of synapse. Since this receptor is genetically associated with schizophrenia, our findings may provide an important clue to understand schizophrenia.

Evaluating Canadians' Values for Drug Coverage Decision Making.

BACKGROUND: Decision makers are facing growing challenges in prioritizing drugs for reimbursement because of soaring drug costs and increasing pressures on financial resources. In addition to cost and effectiveness, payers are using other values to dictate which drugs are prioritized for funding, yet there are limited data on the Canadian public's priorities. OBJECTIVES: To measure the relative societal importance of values considered most relevant in informing drug reimbursement decisions in a representative sample of Canadians. METHODS: An online survey of 2539 Canadians aged 19 years and older was performed in which 13 values used in drug funding prioritization were ranked and then weighted using an analytic hierarchy process. RESULTS: Canadians value safe and efficacious drugs that have certainty of evidence. The values ranked in the top 5 by most of our subjects were potential effect on quality of life (65.4%), severity of the disease (62.6%), ability of drug to work (61.1%), safety (60.5%), and potential to extend life (49.4%). Values related to patient or disease characteristics such as rarity, socioeconomic status, and health and lifestyle choices held the lowest rankings and weights. CONCLUSIONS: Canadians value, above all, treatment-related factors (eg, efficacy and safety) and disease-related factors (eg, severity and equity). Decision makers are currently using additional justifications to prioritize drugs for reimbursement, such as rarity and unmet need, which were not found to be highly valued by Canadians. Decision makers should integrate the public's values into a Canadian reimbursement framework for prioritization of drugs competing for limited funds.

Cardiovascular Risk Reduction in the Workplace With CAMMPUS (Cardiovascular Assessment and Medication Management by Pharmacists at the UBC Site).

BACKGROUND: Cardiovascular (CV) disease is a leading cause of death despite being largely preventable. Employers increasingly offer preventive health programs in the workplace, and pharmacists are well suited to provide these programs. OBJECTIVE: To evaluate the impact of a pharmacist-led service on CV risk in University of British Columbia (UBC) employees. METHODS: This was a prospective observational pre-and-post design study, with participants as their own controls. Employees >18 years of age in the UBC health plan with a Framingham Risk Score (FRS) ≥10% or ≥1 medication-modifiable CV risk factor were included. Participants received a baseline assessment, individualized consultation for 12 months, and a final assessment by a pharmacist at the UBC Pharmacists Clinic. The primary end point was FRS reduction. RESULTS: Baseline assessment of 512 participants between September 2015 and October 2016 yielded 207 (40%) participants, of whom 178 (86%) completed the 12-month intervention. Participants were 54% female and 55% Caucasian, with an average age of 51 (SD = 9.1) years. FRS at baseline was <10 in 45.8%, 10 to 19.9 in 37.9%, and ≥20 in 16.4% of participants. Over 12 months, significant reductions in average FRS (from 11.7 [SD = 7.7] to 10.7 [SD = 7.3]; P = 0.0017) and other parameters were observed. Significant improvements in quality of life (EQ5D change of 0.031 [95% CI = 0.001, 0.062] P = 0.023) and medication adherence (MMAS-8 change of 0.42 [ P = 0.019]) were also noted. CONCLUSIONS AND RELEVANCE: UBC employees had improvements in health markers, self-reported quality of life, and medication adherence after receiving a 12-month pharmacist-led intervention. Pharmacists are encouraged to provide CV risk reduction services in workplaces.

Effect of enhanced homestead food production on anaemia among Cambodian women and children: A cluster randomized controlled trial.

There is inconsistent evidence on the efficacy of agriculture programmes at improving women and children's anaemia and nutritional status. The primary aim of this study was to evaluate the impact of a nutrition-sensitive enhanced homestead food production (EHFP) programme on anaemia in women (18-45 years) and children (6-59 months) in rural Cambodia. Secondary outcomes were women's micronutrient status and women and children's anthropometry. In this cluster-randomized controlled trial, 900 households from 90 villages (clusters) were randomized to either (a) home gardens and behaviour change communication (BCC) on nutrition, hygiene, women's empowerment, and marketing (EHFP); (b) home gardens plus fishponds and BCC (EHFP + F); or (c) control (no intervention). Haemoglobin concentration and anthropometry were measured in women and children at baseline and at 22 months. Venous blood samples were collected in a subset of women (n = 450) at baseline and at 22 months. Generalized linear mixed effect models with repeated measures were used to evaluate the difference across groups and the change from baseline to end of study. Ninety clusters, 552 women, and 754 children completed the trial. Compared with control, we found a statistically significant impact on anaemia prevalence in children (-14.0 percentage points; P = 0.02) and retinol binding protein concentrations in women (difference in difference: 0.34; P = 0.02) randomized to EHFP and EHFP + F groups, respectively. No other statistically significant effects on anaemia, nutritional biomarker concentrations, or anthropometry were observed. Future research is needed to examine longer term impacts of EHFP on anthropometry in women and children and into the nutritional causes of anaemia among children in Cambodia.

Effect of enhanced homestead food production and aquaculture on dietary intakes of women and children in rural Cambodia: A cluster randomized controlled trial.

The Cambodian diet is low in nutrient-dense animal-source foods. Enhanced homestead food production (EHFP) and aquaculture, which increase availability of nutrient-dense foods, are promising interventions to improve dietary intake. This study examined the effect of EHFP with or without aquaculture on dietary intake and prevalence of inadequate intake of select nutrients among women and children living in rural Cambodia, compared to controls. In a registered, cluster randomized controlled trial in Prey Veng, Cambodia, 10 households in each of 90 villages (n = 900) were randomized by village to receive EHFP, EHFP plus aquaculture, or control. After 22-month intervention, 24-hr dietary recalls (24HRs) were collected from mothers aged 18-50 years (n = 429) and their children aged 6 months-7 years (n = 421), reported by their mothers. Usual intake distributions (generated using 24HRs and repeat 24HRs on a subsample) were used to estimate prevalence of inadequate intake. Compared to controls, women in the EHFP group had significantly higher zinc (+1.0 mg/d) and Vitamin A (+139 retinol activity equivalents/d) intakes, and women in the EHFP plus aquaculture group had significantly higher iron (+2.7 mg/d), Vitamin A (+191 retinol activity equivalents/d), and riboflavin (+0.17 mg/d) intakes. Women in the EHFP plus aquaculture group also had significantly lower prevalence of inadequate iron (-7%, at 10% bioavailability), Vitamin A (-19%), and riboflavin (-17%) intakes, compared to controls. No significant differences in intakes or nutrient adequacy were observed among children or between EHFP and EHFP plus aquaculture groups. The biological importance of the small differences in nutrient intakes among women remains to be established.

A non-synonymous single-nucleotide polymorphism associated with multiple sclerosis risk affects the EVI5 interactome.

Despite recent progress in the characterization of genetic loci associated with multiple sclerosis (MS) risk, the ubiquitous linkage disequilibrium operating across the genome has stalled efforts to distinguish causative variants from proxy single-nucleotide polymorphisms (SNPs). Here, we have identified through fine mapping and meta-analysis EVI5 as the most plausible disease risk gene within the 1p22.1 locus. We further show that an exonic SNP associated with risk induces changes in superficial hydrophobicity patterns of the coiled-coil domain of EVI5, which, in turns, affects the EVI5 interactome. Immunoprecipitation of wild-type and mutated EVI5 followed by mass spectrometry generated a roster of disease-specific interactors functionally linked to lipid metabolism. Among the exclusive binding partners of the risk variant, we describe the novel interaction with sphingosine 1-phosphate lyase (SGPL1)-a key enzyme for the creation of the sphingosine-1 phosphate gradient, which is relevant to the pathogenic process and therapeutic management of MS.

A ribonucleoprotein complex protects the interleukin-6 mRNA from degradation by distinct herpesviral endonucleases.

During lytic Kaposi's sarcoma-associated herpesvirus (KSHV) infection, the viral endonuclease SOX promotes widespread degradation of cytoplasmic messenger RNA (mRNA). However, select mRNAs escape SOX-induced cleavage and remain robustly expressed. Prominent among these is interleukin-6 (IL-6), a growth factor important for survival of KSHV infected B cells. IL-6 escape is notable because it contains a sequence within its 3' untranslated region (UTR) that can confer protection when transferred to a SOX-targeted mRNA, and thus overrides the endonuclease targeting mechanism. Here, we pursued how this protective RNA element functions to maintain mRNA stability. Using affinity purification and mass spectrometry, we identified a set of proteins that associate specifically with the protective element. Although multiple proteins contributed to the escape mechanism, depletion of nucleolin (NCL) most severely impacted protection. NCL was re-localized out of the nucleolus during lytic KSHV infection, and its presence in the cytoplasm was required for protection. After loading onto the IL-6 3' UTR, NCL differentially bound to the translation initiation factor eIF4H. Disrupting this interaction, or depleting eIF4H, reinstated SOX targeting of the RNA, suggesting that interactions between proteins bound to distant regions of the mRNA are important for escape. Finally, we found that the IL-6 3' UTR was also protected against mRNA degradation by the vhs endonuclease encoded by herpes simplex virus, despite the fact that its mechanism of mRNA targeting is distinct from SOX. These findings highlight how a multitude of RNA-protein interactions can impact endonuclease targeting, and identify new features underlying the regulation of the IL-6 mRNA.

Syntaxin13 expression is regulated by mammalian target of rapamycin (mTOR) in injured neurons to promote axon regeneration.

Injured peripheral neurons successfully activate intrinsic signaling pathways to enable axon regeneration. We have previously shown that dorsal root ganglia (DRG) neurons activate the mammalian target of rapamycin (mTOR) pathway following injury and that this activity enhances their axon growth capacity. mTOR plays a critical role in protein synthesis, but the mTOR-dependent proteins enhancing the regenerative capacity of DRG neurons remain unknown. To identify proteins whose expression is regulated by injury in an mTOR-dependent manner, we analyzed the protein composition of DRGs from mice in which we genetically activated mTOR and from mice with or without a prior nerve injury. Quantitative label-free mass spectrometry analyses revealed that the injury effects were correlated with mTOR activation. We identified a member of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) family of proteins, syntaxin13, whose expression was increased by injury in an mTOR-dependent manner. Increased syntaxin13 levels in injured nerves resulted from local protein synthesis and not axonal transport. Finally, knockdown of syntaxin13 in cultured DRG neurons prevented axon growth and regeneration. Together, these data suggest that syntaxin13 translation is regulated by mTOR in injured neurons to promote axon regeneration.

Polyglutamine-mediated ribotoxicity disrupts proteostasis and stress responses in Huntington's disease.

Huntington's disease (HD) is a neurodegenerative disorder caused by expansion of a CAG trinucleotide repeat in the Huntingtin (HTT) gene, encoding a homopolymeric polyglutamine (polyQ) tract. Although mutant HTT (mHTT) protein is known to aggregate, the links between aggregation and neurotoxicity remain unclear. Here we show that both translation and aggregation of wild-type HTT and mHTT are regulated by a stress-responsive upstream open reading frame and that polyQ expansions cause abortive translation termination and release of truncated, aggregation-prone mHTT fragments. Notably, we find that mHTT depletes translation elongation factor eIF5A in brains of symptomatic HD mice and cultured HD cells, leading to pervasive ribosome pausing and collisions. Loss of eIF5A disrupts homeostatic controls and impairs recovery from acute stress. Importantly, drugs that inhibit translation initiation reduce premature termination and mitigate this escalating cascade of ribotoxic stress and dysfunction in HD.

Structure and Interactions of HIV-1 gp41 CHR-NHR Reverse Hairpin Constructs Reveal Molecular Determinants of Antiviral Activity.

Engineered reverse hairpin constructs containing a partial C-heptad repeat (CHR) sequence followed by a short loop and full-length N-heptad repeat (NHR) were previously shown to form trimers in solution and to be nanomolar inhibitors of HIV-1 Env mediated fusion. Their target is the in situ gp41 fusion intermediate, and they have similar potency to other previously reported NHR trimers. However, their design implies that the NHR is partially covered by CHR, which would be expected to limit potency. An exposed hydrophobic pocket in the folded structure may be sufficient to confer the observed potency, or they may exist in a partially unfolded state exposing full length NHR. Here we examined their structure by crystallography, CD and fluorescence, establishing that the proteins are folded hairpins both in crystal form and in solution. We examined unfolding in the milieu of the fusion reaction by conducting experiments in the presence of a membrane mimetic solvent and by engineering a disulfide bond into the structure to prevent partial unfolding. We further examined the role of the hydrophobic pocket, using a hairpin-small molecule adduct that occluded the pocket, as confirmed by X-ray footprinting. The results demonstrated that the NHR region nominally covered by CHR in the engineered constructs and the hydrophobic pocket region that is exposed by design were both essential for nanomolar potency and that interaction with membrane is likely to play a role in promoting the required inhibitor structure. The design concepts can be applied to other Class 1 viral fusion proteins.

Contrastsing synaptic roles of MDGA1 and MDGA2.

Neurodevelopmental disorders are frequently linked to mutations in synaptic organizing molecules. MAM domain containing glycosylphosphatidylinositol anchor 1 and 2 (MDGA1 and MDGA2) are a family of synaptic organizers suggested to play an unusual role as synaptic repressors, but studies offer conflicting evidence for their localization. Using epitope-tagged MDGA1 and MDGA2 knock-in mice, we found that native MDGAs are expressed throughout the brain, peaking early in postnatal development. Surprisingly, endogenous MDGA1 was enriched at excitatory, but not inhibitory, synapses. Both shRNA knockdown and CRISPR/Cas9 knockout of MDGA1 resulted in cell-autonomous, specific impairment of AMPA receptor-mediated synaptic transmission, without affecting GABAergic transmission. Conversely, MDGA2 knockdown/knockout selectively depressed NMDA receptor-mediated transmission but enhanced inhibitory transmission. Our results establish that MDGA2 acts as a synaptic repressor, but only at inhibitory synapses, whereas both MDGAs are required for excitatory transmission. This nonoverlapping division of labor between two highly conserved synaptic proteins is unprecedented.

A Legionella toxin exhibits tRNA mimicry and glycosyl transferase activity to target the translation machinery and trigger a ribotoxic stress response.

A widespread strategy employed by pathogens to establish infection is to inhibit host-cell protein synthesis. Legionella pneumophila, an intracellular bacterial pathogen and the causative organism of Legionnaires' disease, secretes a subset of protein effectors into host cells that inhibit translation elongation. Mechanistic insights into how the bacterium targets translation elongation remain poorly defined. We report here that the Legionella effector SidI functions in an unprecedented way as a transfer-RNA mimic that directly binds to and glycosylates the ribosome. The 3.1 Å cryo-electron microscopy structure of SidI reveals an N-terminal domain with an 'inverted L' shape and surface-charge distribution characteristic of tRNA mimicry, and a C-terminal domain that adopts a glycosyl transferase fold that licenses SidI to utilize GDP-mannose as a sugar precursor. This coupling of tRNA mimicry and enzymatic action endows SidI with the ability to block protein synthesis with a potency comparable to ricin, one of the most powerful toxins known. In Legionella-infected cells, the translational pausing activated by SidI elicits a stress response signature mimicking the ribotoxic stress response, which is activated by elongation inhibitors that induce ribosome collisions. SidI-mediated effects on the ribosome activate the stress kinases ZAKα and p38, which in turn drive an accumulation of the protein activating transcription factor 3 (ATF3). Intriguingly, ATF3 escapes the translation block imposed by SidI, translocates to the nucleus and orchestrates the transcription of stress-inducible genes that promote cell death, revealing a major role for ATF3 in the response to collided ribosome stress. Together, our findings elucidate a novel mechanism by which a pathogenic bacterium employs tRNA mimicry to hijack a ribosome-to-nuclear signalling pathway that regulates cell fate.