Maternally supplied S-acyl-transferase is required for crystalloid organelle formation and transmission of the malaria parasite.

Transmission of the malaria parasite from the mammalian host to the mosquito vector requires the formation of adequately adapted parasite forms and stage-specific organelles. Here we show that formation of the crystalloid-a unique and short-lived organelle of the Plasmodium ookinete and oocyst stage required for sporogony-is dependent on the precisely timed expression of the S-acyl-transferase DHHC10. DHHC10, translationally repressed in female Plasmodium berghei gametocytes, is activated translationally during ookinete formation, where the protein is essential for the formation of the crystalloid, the correct targeting of crystalloid-resident protein LAP2, and malaria parasite transmission

Skin Autofluorescence and Pentosidine Are Associated With Aortic Stiffening:The Maastricht Study

Arterial stiffening, as characterized by an increase in carotid-femoral pulse-wave velocity or pulse pressure, increases the risk of cardiovascular disease, especially among individuals with type 2 diabetes mellitus. Advanced glycation end products are hypothesized to play a role in the development of arterial stiffness. Therefore, we investigated the association between skin autofluorescence, an estimate of tissue advanced glycation end products, and plasma advanced glycation end products on the one hand and arterial stiffening on the other in 862 participants of The Maastricht Study (mean age of 60 years; 45% women) with normal glucose metabolism (n=469), impaired glucose metabolism (n=140), or type 2 diabetes (n=253). Associations were analyzed with linear regression analysis and adjusted for potential confounders. We found that higher skin autofluorescence as measured by the AGE Reader and plasma pentosidine were independently associated with higher carotid-femoral pulse-wave velocity (s 0.10; 95% confidence interval, 0.03-0.17 and 0.10; 0.04-0.16, respectively) and central pulse pressure (s 0.08; 95% confidence interval 0.01-0.15 and 0.07; 0.01-0.13, respectively). The associations between skin autofluorescence and pentosidine, and carotid-femoral pulse-wave velocity were more pronounced in individuals with type 2 diabetes mellitus (P-interaction</p

Small cargo proteins and large aggregates can traverse the Golgi by a common mechanism without leaving the lumen of cisternae

Procollagen (PC)-I aggregates transit through the Golgi complex without leaving the lumen of Golgi cisternae. Based on this evidence, we have proposed that PC-I is transported across the Golgi stacks by the cisternal maturation process. However, most secretory cargoes are small, freely diffusing proteins, thus raising the issue whether they move by a transport mechanism different than that used by PC-I. To address this question we have developed procedures to compare the transport of a small protein, the G protein of the vesicular stomatitis virus (VSVG), with that of the much larger PC-I aggregates in the same cell. Transport was followed using a combination of video and EM, providing high resolution in time and space. Our results reveal that PC-I aggregates and VSVG move synchronously through the Golgi at indistinguishable rapid rates. Additionally, not only PC-I aggregates (as confirmed by ultrarapid cryofixation), but also VSVG, can traverse the stack without leaving the cisternal lumen and without entering Golgi vesicles in functionally relevant amounts. Our findings indicate that a common mechanism independent of anterograde dissociative carriers is responsible for the traffic of small and large secretory cargo across the Golgi stack

Optimisations and challenges involved in the creation of various bioluminescent and fluorescent influenza a virus strains for in vitro and in vivo applications

Bioluminescent and fluorescent influenza A viruses offer new opportunities to study influenza virus replication, tropism and pathogenesis. To date, several influenza A reporter viruses have been described. These strategies typically focused on a single reporter gene (either bioluminescent or fluorescent) in a single virus backbone. However, whilst bioluminescence is suited to in vivo imaging, fluorescent viruses are more appropriate for microscopy. Therefore, the idea l reporter virus varies depending on the experiment in question, and it is important that any reporter virus strategy can be adapted accordingly. Herein, a strategy was developed to create five different reporter viruses in a single virus backbone. Specifically, enhanced green fluorescent protein (eGFP), far-red fluorescent protein (fRFP), near-infrared fluorescent protein (iRFP), Gaussia luciferase (gLUC) and firefly luciferase (fLUC) were inserted into the PA gene segment of A/PR/8/34 (H1N1). This study provides a comprehensive characterisation of the effects of different reporter genes on influenza virus replication and reporter activity. In vivo reporter gene expression, in lung tissues, was only detected for eGFP, fRFP and gLUC expressing viruses. In vitro, the eGFP-expressing virus displayed the best reporter stability and could be used for correlative light electron microscopy (CLEM). This strategy was then used to create eGFP-expressing viruses consisting entirely of pandemic H1N1, highly pathogenic avian influenza (HPAI) H5N1 and H7N9. The HPAI H5N1 eGFP-expressing virus infected mice and reporter gene expression was detected, in lung tissues, in vivo. Thus, this study provides new tools and insights for the creation of bioluminescent and fluorescent influenza A reporter viruses. Copyright

Prediabetes Is Associated With Structural Brain Abnormalities:The Maastricht Study

OBJECTIVE Structural brain abnormalities are key risk factors for brain diseases, such as dementia, stroke, and depression, in type 2 diabetes. It is unknown whether structural brain abnormalities already occur in prediabetes. Therefore, we investigated whether both prediabetes and type 2 diabetes are associated with lacunar infarcts (LIs), white matter hyperintensities (WMHs), cerebral microbleeds (CMBs), and brain atrophy. RESEARCH DESIGN and METHODS We used data from 2,228 participants (1,373 with normal glucose metabolism [NGM], 347 with prediabetes, and 508 with type 2 diabetes (oversampled); mean age 59.2 6 8.2 years; 48.3% women) of the Maastricht Study, a population-based cohort study. Diabetes status was determined with an oral glucose tolerance test. Brain imaging was performed with 3 Tesla MRI. Results were analyzed with multivariable logistic and linear regression analyses. RESULTS Prediabetes and type 2 diabetes were associated with the presence of LIs (odds ratio 1.61 [95% CI 0.98-2.63] and 1.67 [1.04-2.68], respectively; P trend = 0.027), larger WMH (b 0.07 log10-transformed mL [log-mL] [95% CI 0.00-0.15] and 0.21 log-mL [0.14-0.28], respectively; P trend <0.001), and smaller white matter volumes (b 24.0 mL [27.3 to 20.6] and 27.2 mL [210.4 to 24.0], respectively; P trend <0.001) compared with NGM. Prediabetes was not associated with gray matter volumes or the presence of CMBs. CONCLUSIONS Prediabetes is associated with structural brain abnormalities, with further deterioration in type 2 diabetes. These results indicate that, in middle-aged populations, structural brain abnormalities already occur in prediabetes, which may suggest that the treatment of early dysglycemia may contribute to the prevention of brain diseases

Estimated GFR, Albuminuria, and Cognitive Performance:The Maastricht Study

BACKGROUND: Reduced estimated glomerular filtration rate (eGFR) and albuminuria have been associated with worse cognitive performance. However, few studies have examined whether these associations are confined to older individuals or may be extended to the middle-aged population. STUDY DESIGN: Cross-sectional analyses of a prospective population-based cohort study. SETTING & PARTICIPANTS: 2,987 individuals aged 40 to 75 years from the general population (The Maastricht Study). PREDICTOR: eGFR and urinary albumin excretion (UAE). OUTCOMES: Memory function, information processing speed, and executive function. MEASUREMENTS: Analyses were adjusted for demographic variables (age, sex, and educational level), lifestyle factors (smoking behavior and alcohol consumption), depression, and cardiovascular disease risk factors (glucose metabolism status, waist circumference, total to high-density lipoprotein cholesterol ratio, triglyceride level, use of lipid-modifying medication, systolic blood pressure, use of antihypertensive medication, and prevalent cardiovascular disease). RESULTS: UAE was <15mg/24 h in 2,439 (81.7%) participants, 15 to <30 mg/24 h in 309 (10.3%), and ≥30mg/24 h in 239 (8.0%). In the entire study population, UAE≥30mg/24 h was associated with lower information processing speed as compared to UAE<15mg/24 h (β [SD difference] = -0.148; 95% CI, -0.263 to -0.033) after full adjustment, whereas continuous albuminuria was not. However, significant interaction terms (P for interaction < 0.05) suggested that albuminuria was most strongly and extensively associated with cognitive performance in older individuals. Mean (±SD) eGFR, estimated by the CKD-EPI (Chronic Kidney Disease Epidemiology Collaboration) creatinine-cystatin C equation (eGFRcr-cys), was 88.4±14.6 mL/min/1.73m(2). eGFRcr-cys was not associated with any of the domains of cognitive performance after full adjustment. However, significant interaction terms (P for interaction < 0.05) suggested that eGFRcr-cys was associated with cognitive performance in older individuals. LIMITATIONS: Cross-sectional design, which limited causal inferences. CONCLUSIONS: In the entire study population, albuminuria was independently associated with lower information processing speed, whereas eGFRcr-cys was not associated with cognitive performance. However, both were more strongly and extensively associated with cognitive performance in older individuals

Multiple capsid-stabilizing interactions revealed in a high-resolution structure of an emerging picornavirus causing neonatal sepsis

The poorly studied picornavirus, human parechovirus 3 (HPeV3) causes neonatal sepsis with no therapies available. Our 4.3-Å resolution structure of HPeV3 on its own and at 15 Å resolution in complex with human monoclonal antibody Fabs demonstrates the expected picornavirus capsid structure with three distinct features. First, 25% of the HPeV3 RNA genome in 60 sites is highly ordered as confirmed by asymmetric reconstruction, and interacts with conserved regions of the capsid proteins VP1 and VP3. Second, the VP0 N terminus stabilizes the capsid inner surface, in contrast to other picornaviruses where on expulsion as VP4, it forms an RNA translocation channel. Last, VP1's hydrophobic pocket, the binding site for the antipicornaviral drug, pleconaril, is blocked and thus inappropriate for antiviral development. Together, these results suggest a direction for development of neutralizing antibodies, antiviral drugs based on targeting the RNA-protein interactions and dissection of virus assembly on the basis of RNA nucleation.Peer reviewe

Association of Type 2 Diabetes, According to the Number of Risk Factors Within Target Range, With Structural Brain Abnormalities, Cognitive Performance, and Risk of Dementia

OBJECTIVE: Type 2 diabetes is associated with increased risks of cognitive dysfunction and brain abnormalities. The extent to which risk factor modification can mitigate these risks is unclear. We investigated the associations between incident dementia, cognitive performance, and brain abnormalities among individuals with type 2 diabetes, according to the number of risk factors on target, compared with control subjects without diabetes. RESEARCH DESIGN AND METHODS: Prospective data were from UK Biobank of 87,856 individuals (n = 10,663 diabetes, n = 77,193 control subjects; baseline 2006-2010), with dementia follow-up until February 2018. Individuals with diabetes were categorized according to the number of seven selected risk factors within the guideline-recommended target range (nonsmoking; guideline-recommended levels of glycated hemoglobin, blood pressure, BMI, albuminuria, physical activity, and diet). Outcomes were incident dementia, domain-specific cognitive performance, white matter hyperintensities, and total brain volume. RESULTS: After a mean follow-up of 9.0 years, 147 individuals (1.4%) with diabetes and 412 control subjects (0.5%) had incident dementia. Among individuals with diabetes, excess dementia risk decreased stepwise for a higher number of risk factors on target. Compared with control subjects (incidence rate per 1,000 person-years 0.62 [95% CI 0.56; 0.68]), individuals with diabetes who had five to seven risk factors on target had no significant excess dementia risk (absolute rate difference per 1,000 person-years 0.20 [-0.11; 0.52]; hazard ratio 1.32 [0.89; 1.95]). Similarly, differences in processing speed, executive function, and brain volumes were progressively smaller for a higher number of risk factors on target. These results were replicated in the Maastricht Study. CONCLUSIONS: Among individuals with diabetes, excess dementia risk, lower cognitive performance, and brain abnormalities decreased stepwise for a higher number of risk factors on target

SARS-Coronavirus Replication Is Supported by a Reticulovesicular Network of Modified Endoplasmic Reticulum

Positive-strand RNA viruses, a large group including human pathogens such as SARS-coronavirus (SARS-CoV), replicate in the cytoplasm of infected host cells. Their replication complexes are commonly associated with modified host cell membranes. Membrane structures supporting viral RNA synthesis range from distinct spherular membrane invaginations to more elaborate webs of packed membranes and vesicles. Generally, their ultrastructure, morphogenesis, and exact role in viral replication remain to be defined. Poorly characterized double-membrane vesicles (DMVs) were previously implicated in SARS-CoV RNA synthesis. We have now applied electron tomography of cryofixed infected cells for the three-dimensional imaging of coronavirus-induced membrane alterations at high resolution. Our analysis defines a unique reticulovesicular network of modified endoplasmic reticulum that integrates convoluted membranes, numerous interconnected DMVs (diameter 200–300 nm), and “vesicle packets” apparently arising from DMV merger. The convoluted membranes were most abundantly immunolabeled for viral replicase subunits. However, double-stranded RNA, presumably revealing the site of viral RNA synthesis, mainly localized to the DMV interior. Since we could not discern a connection between DMV interior and cytosol, our analysis raises several questions about the mechanism of DMV formation and the actual site of SARS-CoV RNA synthesis. Our data document the extensive virus-induced reorganization of host cell membranes into a network that is used to organize viral replication and possibly hide replicating RNA from antiviral defense mechanisms. Together with biochemical studies of the viral enzyme complex, our ultrastructural description of this “replication network” will aid to further dissect the early stages of the coronavirus life cycle and its virus-host interactions