Search

C. elegansis a free-living nematode that must adapt to a wide range of environments including both aerobic and anaerobic conditions. To survive in low oxygen,C. eleganscan use an unusual form of anaerobic respiration that relies on rhodoquinone (RQ) as an alternative electron carrier. Parasitic nematodes like hookworm and whipworm also require rhodoquinone-dependent metabolism (RQDM) to survive in the highly anaerobic conditions in the human gut. Understanding how RQDM is regulated inC. elegansmay thus identify new ways to combat these closely-related major human pathogens. We previously established a simple movement-based assay for RQDM inC. elegans. In this study, we tested a panel of wild-type isolates ofC. elegansin our RQDM assay and find substantial variation in their ability to use RQDM. We carried out a genome-wide association study (GWAS) to identify loci that affect RQDM — this identified a single major QTL on the right arm of Chromosome III. We used RNAi to test almost all genes within the QTL region for involvement in RQDM and found one gene,wah-1, that strongly modulates RQDM-dependent recovery inC. elegans. WAH-1 is a mitochondrial flavoprotein that affects the electron transport chain, consistent with a role in RQDM. We show thatwah-1expression varies between isolates due to major changes inwah-1transcript structures and this correlates tightly with variation in RQDM. Finally, we show that there is similar complexity towah-1transcription in parasitic nematodes and thatwah-1transcript structures change as parasites shift from aerobic to anaerobic, RQ- requiring metabolism. We thus conclude that reducedwah-1expression correlates with increased ability to survive in conditions where RQDM is essential.

Soil transmitted helminths (STHs) are major human pathogens that infect over a billion people. Resistance to current anthelmintics is rising and new drugs are needed. Here we combine multiple approaches to find druggable targets that are essential for RQ-dependent metabolism, an unusual form of anaerobic metabolism which STHs need to survive in their host. We identified 25 genes predicted to act in RQ-dependent metabolism from sensing hypoxia to RQ synthesis — this includes components of the kynurenine pathway we previously showed to be essential for RQ synthesis (Del Borrello et al., 2019). We found 9 genes to be required — since all have host orthologues, we used comparative genomics and structural modeling to identify those with helminth-specific active sites and found 4 such targets. These 4 high confidence targets open up the possibility of in silico screens to identify STH-specific inhibitors of these enzymes as new anthelmintics.

Parasitic helminths use two benzoquinones as electron carriers in the electron transport chain. In normoxia, they use ubiquinone (UQ), but in anaerobic conditions inside the host, they require rhodoquinone (RQ) and greatly increase RQ levels. We previously showed the switch from UQ to RQ synthesis is driven by a change of substrates by the polyprenyltransferase COQ-2 (Del Borrello et al., 2019; Roberts Buceta et al., 2019); however, the mechanism of substrate selection is not known. Here, we show helminths synthesize twocoq-2splice forms,coq-2aandcoq-2e, and thecoq-2e-specific exon is only found in species that synthesize RQ. We show that inCaenorhabditis elegansCOQ-2e is required for efficient RQ synthesis and survival in cyanide. Importantly, parasites switch from COQ-2a to COQ-2e as they transit into anaerobic environments. We conclude helminths switch from UQ to RQ synthesis principally via changes in the alternative splicing ofcoq-2.

Parasitic helminths use two benzoquinones as electron carriers in the electron transport chain. In normoxia, they use ubiquinone (UQ), but in anaerobic conditions inside the host, they require rhodoquinone (RQ) and greatly increase RQ levels. We previously showed the switch from UQ to RQ synthesis is driven by a change of substrates by the polyprenyltransferase COQ-2 (Del Borrello et al., 2019; Roberts Buceta et al., 2019); however, the mechanism of substrate selection is not known. Here, we show helminths synthesize two coq-2 splice forms, coq-2a and coq-2e, and the coq-2e-specific exon is only found in species that synthesize RQ. We show that in Caenorhabditis elegans COQ-2e is required for efficient RQ synthesis and survival in cyanide. Importantly, parasites switch from COQ-2a to COQ-2e as they transit into anaerobic environments. We conclude helminths switch from UQ to RQ synthesis principally via changes in the alternative splicing of coq-2.

Parasitic helminths use two benzoquinones as electron carriers in the electron transport chain. In aerobic environments they use ubiquinone (UQ) but in anaerobic environments inside the host, they require rhodoquinone (RQ) and greatly increase RQ levels. The switch to RQ synthesis is driven by substrate selection by the polyprenyltransferase COQ-2 but the mechanisms underlying this substrate choice are unknown. We found that helminths make twocoq-2isoforms,coq-2aandcoq-2e, by alternative splicing. COQ-2a is homologous to COQ2 from other eukaryotes but the COQ-2e-specific exon is only found in species that make RQ and its inclusion changes the enzyme core. We show COQ-2e is required for RQ synthesis and for survival in cyanide inC. elegans. Crucially, we see a switch from COQ-2a to COQ-2e as parasites transition into anaerobic environments. We conclude that under anaerobic conditions helminths switch from UQ to RQ synthesis via alternative splicing ofcoq-2.

Parasitic helminths infect over a billion humans. To survive in the low oxygen environment of their hosts, these parasites use unusual anaerobic metabolism — this requires rhodoquinone (RQ), an electron carrier that is made by very few animal species. Crucially RQ is not made or used by any parasitic hosts and RQ synthesis is thus an ideal target for anthelmintics. However, little is known about how RQ is made and no drugs are known to block RQ synthesis. C. elegans makes RQ and can use RQ-dependent metabolic pathways — here, we use C. elegans genetics to show that tryptophan degradation via the kynurenine pathway is required to generate the key amine-containing precursors for RQ synthesis. We show that C. elegans requires RQ for survival in hypoxic conditions and, finally, we establish a high throughput assay for drugs that block RQ-dependent metabolism. This may drive the development of a new class of anthelmintic drugs. This study is a key first step in understanding how RQ is made in parasitic helminths., eLife digest Parasitic worms infect more than a billion people worldwide, using a range of tricks to survive inside the human body. Some species can live for weeks inside the gut, a place with almost no oxygen. Yet exactly how they manage this is remains unclear. Scientists know that parasitic worms have an unusual way of making chemical energy when oxygen levels drop. Like human cells, worm cells use a series of molecular complexes called the electron transport chain. As electrons pass along the chain, they drive the production of chemical energy. Normally, oxygen sits at the end of the chain to receive the electrons. But, when there is no oxygen, almost all animals stop using the electron transport chain. A few animals can continue to use it by using other molecules to receive the final electrons instead of oxygen. To do that, they need a special electron carrier and, in worms, this electron carrier is rhodoquinone. Human cells do not use rhodoquinone, making it a prime target for drug design. If a drug could block rhodoquinone production, it might be able to stop worms surviving in the human intestines without harming the patient’s own cells. Yet, even though the scientific community has known about rhodoquinone for more than 50 years, it remains unclear how worms make this molecule. To find out, Del Borrello et al. examined the laboratory worm Caenorhabditis elegans. This worm is not a parasite, but it does make rhodoquinone. Del Borrello et al. developed a new way to study rhodoquinone production by blocking the normal route of the electron transport chain with cyanide. This causes the worms to switch to using rhodoquinone and is cheaper than raising the worms in low oxygen, making it easier to conduct high-throughput screening. A combination of chemistry and information from other species made it possible to identify candidate genes responsible for the production of rhodoquinone. Worms with faults in these genes revealed the key building blocks of rhodoquinone, and the early steps in its production. Removing any one of the genes made it harder for the worms to survive without oxygen. Although there are already effective drugs that kill parasitic worms, resistance is growing. A better understanding of rhodoquinone could lead to a new class of drugs to help control this major problem in global health. A drug that blocks any one of the production steps of rhodoquinone might be a future candidate for a new anti-parasitic worm therapy.

Parasitic helminths infect over a billion humans. To survive in the low oxygen environment of their hosts, these parasites use unusual anaerobic metabolism. This requires Rhodoquinone (RQ), an electron carrier that is made by very few animal species — crucially it is not present in any parasitic hosts. RQ synthesis is thus an ideal target for anthelmintics but little is known about how RQ is made and no drugs are known to block RQ synthesis.C.elegansmakes RQ and can use RQ-dependent metabolic pathways — here, we useC.elegansgenetics to identify the pathway for RQ synthesis and show thatC.elegansrequires RQ for survival in hypoxic conditions. Finally, we establish a robust assay for drugs that block RQ-dependent metabolism. This study identifies for the first time how RQ is made in any animal and establishes a novel assay that can drive the development of a new class of anthelmintic drugs.

Protein complexes are key macromolecular machines of the cell, but their description remains incomplete. We and others previously reported an experimental strategy for global characterization of native protein assemblies based on chromatographic fractionation of biological extracts coupled to precision mass spectrometry analysis (chromatographic fractionation-mass spectrometry, CF-MS), but the resulting data are challenging to process and interpret. Here, we describe EPIC (elution profile-based inference of complexes), a software toolkit for automated scoring of large-scale CF-MS data to define high-confidence multi-component macromolecules from diverse biological specimens. As a case study, we used EPIC to map the global interactome of Caenorhabditis elegans, defining 612 putative worm protein complexes linked to diverse biological processes. These included novel subunits and assemblies unique to nematodes that we validated using orthogonal methods. The open source EPIC software is freely available as a Jupyter notebook packaged in a Docker container (https://hub.docker.com/r/baderlab/bio-epic/).

Treatment rates with intravenously administered tissue plasminogen activator (IV-tPA) in acute ischemic stroke (IS) remain low in Asian populations. Various logistic obstacles and higher anticipated bleeding risk in Asians are major concerns. We report on the feasibility and safety of IV-tPA therapy at our tertiary care center. Consecutive acute IS patients eligible for thrombolysis were treated with low-dose (maximum 50 mg) IV-tPA between January 2000 and September 2006 and with standard-dose (maximum 90 mg) IV-tPA between October 2006 and May 2008. The efficacy of IV-tPA was assessed by the modified Rankin Scale (mRS) score at 3 months and by absolute changes in the National Institute of Health Stroke Scale (NIHSS) score at hospital discharge and 3 months. The safety of IV-tPA was assessed by the rate of symptomatic intracranial hemorrhage (SICH). A total of 130 patients were included (mean age, 60±13 years; 60% males; median NIHSS score, 14). A total of 48 patients received low-dose IV-tPA, and 82 patients received standard-dose IV-tPA. The median onset to treatment time was 160 minutes. Some 59% of the patients achieved functional independence (mRS score 0-1) at 3 months with standard-dose tPA, compared with 35% in the low-dose group (P=.011). SICH occurred more frequently with the low dose (14.5%) than with the standard dose (1.2%; P=.004). In a multivariate logistic regression model, lower admission NIHSS score (odds ratio [OR]=0.78 per 1-point increase; 95% confidence interval [CI]=0.70-0.88), lower pretreatment blood glucose level (OR=0.76 per 1 mmol/L increase; 95% CI=0.60-0.95), shorter time from symptom onset to IV-tPA bolus (OR=0.97 per 1-minute increase; 95% CI=0.94-1.0), and standard-dose IV-tPA (OR=12.49; 95% CI=2.9-53.89) were associated with a higher likelihood for functional independence at 3 months. Our data indicate that standard-dose IV-tPA (0.9 mg/kg) was feasible and safe for treating acute IS in our multiethnic Asian population in Singapore.

Objective: To describe the patterns of early and delayed hypoxic damage to the brain parenchyma.Methods: This paper reports a case with interesting clinical course and MRI brain changes in a patient following a single hypoxic-anoxic exposure after an alcoholic binge.Results: Bilaterally symmetrical putaminal necrosis was noted in the initial MRI, but subsequent follow-up revealed extensive demyelination of the sub-cortical white matter, associated with the clinical deterioration.Conclusions: The brain, in general, and the neurons in caudate nucleus, putamen, globus pallidus and cerebral white matter, in particular, are susceptible to hypoxic injury. These changes may occur simultaneously and early or sequentially and late.

Background Prospective studies on lipids and risk of Parkinson9s disease (PD) in Asian populations are sparse. This study prospectively examined the associations between dietary cholesterol and major fatty acids, and risk of PD among the Chinese in Singapore. Methods This study used data from the Singapore Chinese Health Study, a population-based prospective cohort of 63 257 men and women aged 45–74 years in Singapore enrolled in 1993–1998. Dietary intakes of cholesterol and fatty acids were derived from a validated semiquantitative food frequency questionnaire and the Singapore Food Composition Table. Incident PD cases were identified either through follow-up interviews or record linkage analysis with hospital discharge and PD outpatient registries. Results After an average of 14.6 years, 218 men and 193 women in the cohort developed PD. Dietary cholesterol was associated with statistically significantly lower risk of PD in a dose–dependent manner among men after adjustment for established risk factors for PD and intakes of major fatty acids. Compared to the lowest quartile, HR (95% CI) for the highest quartile was 0.53 (95% CI 0.33 to 0.84) (P for trend=0.006). Among women, dietary monounsaturated fatty acid was inversely associated with PD risk (P for trend=0.033). Compared to the lowest quartile, HR for the highest quartile was 0.44 (95% CI 0.22 to 0.88). There was no statistically significant association between dietary saturated, n-3 and n-6 fatty acids and PD risk. Conclusions Higher intakes of cholesterol and monounsaturated fatty acids may reduce risk of PD in men and women, respectively.

Data from Asian populations on dietary and lifestyle factors associated with Parkinson's disease are sparse. In 1993-2005, the authors examined these factors in relation to Parkinson's disease in the Singapore Chinese Health Study, a prospective cohort of 63,257 Chinese men and women. Baseline data were collected through in-person interviews using structured questionnaires. All 157 incident Parkinson's disease cases were identified either through follow-up interviews or via linkage with hospital discharge databases and Parkinson's disease outpatient registries and were confirmed by review of medical records. Current versus never smokers exhibited a reduced risk of Parkinson's disease (relative risk = 0.29, 95% confidence interval: 0.16, 0.52). Total caffeine intake was inversely related to Parkinson's disease risk (p for trend = 0.002); the relative risk for the highest versus lowest quartile was 0.55 (95% confidence interval: 0.35, 0.88). Black tea, a caffeine-containing beverage, showed an inverse association with Parkinson's disease risk that was not confounded by total caffeine intake or tobacco smoking (p for trend = 0.0006; adjusted relative risk for the highest vs. lowest tertile of intake = 0.29, 95% confidence interval: 0.13, 0.67). Green tea drinking was unrelated to Parkinson's disease risk. Diet had no strong influence on risk. Ingredients of black tea other than caffeine appear to be responsible for the beverage's inverse association with Parkinson's disease.