Proteomic analysis of lipid raft-enriched membranes isolated from internal organelles

The mitochondria-associated membrane (MAM) is a sub-region of the endoplasmic reticulum (ER) that facilitates crosstalk between the ER and mitochondria. The MAM actively influences vital cellular processes including Ca²⁺ signaling and protein folding. Detergent-resistant microdomains (DRMs) may localize proteins to the mitochondria/MAM interface to coordinate these events. However, the protein composition of DRMs isolated from this region is not known. Lipid-raft enriched DRMs were isolated from a combined mitochondria/MAM sample and analyzed using two-dimensional reversed-phased tandem mass spectrometry. Strict post-acquisition filtering of the acquired data led to the confident identification 250 DRM proteins. The majority (58%) of the identified proteins are bona fide mitochondrial or ER proteins according to Gene Ontology annotation. Additionally, 74% of the proteins have previously been noted as MAM-resident or -associated proteins. Furthermore, ∼20% of the identified proteins have a documented association with lipid rafts. Most importantly, known internal LR marker proteins (inositol 1,4,5-trisphosphate receptor type 3, erlin-2, and voltage-dependent anion channel 1) were detected as well as most of the components of the mitochondrial/MAM-localized Ca²⁺ signaling complex. Our study provides the basis for future work probing how the protein activities at the mitochondrion/MAM interface are dependent upon the integrity of these internal lipid-raft-like domains.     

FANCM-associated proteins MHF1 and MHF2, but not the other Fanconi anemia factors,limit meiotic crossovers

Genetic recombination is important for generating diversity and to ensure faithful segregation of chromosomes at meiosis. However, few crossovers (COs) are formed per meiosis despite an excess of DNA double-strand break precursors. This reflects the existence of active mechanisms that limit CO formation. We previously showed that AtFANCM is a meiotic anti-CO factor. The same genetic screen now identified AtMHF2 as another player of the same anti-CO pathway. FANCM and MHF2 are both Fanconi Anemia (FA) associated proteins, prompting us to test the other FA genes conserved in Arabidopsis for a role in CO control at meiosis. This revealed that among the FA proteins tested, only FANCM and its two DNA-binding co-factors MHF1 and MHF2 limit CO formation at meiosis.     

PPARγ Agonists Improve Survival and Neurocognitive Outcomes in Experimental Cerebral Malaria and Induce Neuroprotective Pathways in Human Malaria

Cerebral malaria (CM) is associated with a high mortality rate, and long-term neurocognitive impairment in approximately one third of survivors. Adjunctive therapies that modify the pathophysiological processes involved in CM may improve outcome over anti-malarial therapy alone. PPARγ agonists have been reported to have immunomodulatory effects in a variety of disease models. Here we report that adjunctive therapy with PPARγ agonists improved survival and long-term neurocognitive outcomes in the Plasmodium berghei ANKA experimental model of CM. Compared to anti-malarial therapy alone, PPARγ adjunctive therapy administered to mice at the onset of CM signs, was associated with reduced endothelial activation, and enhanced expression of the anti-oxidant enzymes SOD-1 and catalase and the neurotrophic factors brain derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in the brains of infected mice. Two months following infection, mice that were treated with anti-malarials alone demonstrated cognitive dysfunction, while mice that received PPARγ adjunctive therapy were completely protected from neurocognitive impairment and from PbA-infection induced brain atrophy. In humans with P. falciparum malaria, PPARγ therapy was associated with reduced endothelial activation and with induction of neuroprotective pathways, such as BDNF. These findings provide insight into mechanisms conferring improved survival and preventing neurocognitive injury in CM, and support the evaluation of PPARγ agonists in human CM.     

Determinants of Mortality and Loss to Follow-Up among Adults Enrolled in HIV Care Services in Rwanda

Background

Antiretroviral therapy (ART) improves morbidity and mortality in patients with HIV, however high rates of loss to follow-up (LTF) and mortality have been documented in HIV care and treatment programs.

Methods

We analyzed routinely-collected data on HIV-infected patients ≥15 years enrolled at 41 healthcare facilities in Rwanda from 2005 to 2010. LTF was defined as not attending clinic in the last 12 months for pre-ART patients and 6 months for ART patients. For the pre-ART period, sub-distribution hazards models were constructed to estimate LTF and death to account for competing risks. Kaplan-Meier (KM) and Cox proportional hazards models were used for patients on ART.

Results

31,033 ART-naïve adults were included, 64% were female and 75% were WHO stage I or II at enrollment. 17,569 (56%) patients initiated ART. Pre-ART competing risk estimates of LTF at 2 years was 11.2% (95%CI, 10.9–11.6%) and 2.9% for death (95%CI 2.7–3.1%). Among pre-ART patients, male gender was associated with higher LTF (adjusted sub-hazard ratio (aSHR) 1.3, 95%CI 1.1–1.5) and death (aSHR 1.7, 95%CI 1.4–2.1). Low CD4 count (CD4<100 vs. ≥350 aSHR 0.2, 95%CI 0.1–0.3) and higher WHO stage (WHO stage IV vs. stage I aSHR 0.4, 95%CI 0.2–0.6) were protective against pre-ART LTF. KM estimates for LTF and death in ART patients at 2 years were 4.4% (95%CI 4.4–4.5%) and 6.3% (95%CI 6.2–6.4%). In patients on ART, male gender was associated with LTF (adjusted hazard ratio (AHR) 1.4, 95%CI 1.2–1.7) and death (AHR1.3, 95%CI 1.2–1.5). Mortality was higher for ART patients ≥40 years and in those with lower CD4 count at ART initiation.

Conclusions

Low rates of LTF and death were founds among pre-ART and ART patients in Rwanda but greater efforts are needed to retain patients in care prior to ART initiation, particularly among those who are healthy at enrollment.     

Pre-Existing Hypertension Dominates γδT Cell Reduction in Human Ischemic Stroke

T lymphocytes may play an important role in the evolution of ischemic stroke. Depletion of γδT cells has been found to abrogate ischemia reperfusion injury in murine stroke. However, the role of γδT cells in human ischemic stroke is unknown. We aimed to determine γδT cell counts and γδT cell interleukin 17A (IL-17A) production in the clinical setting of ischemic stroke. We also aimed to determine the associations of γδT cell counts with ischemic lesion volume, measures of clinical severity and with major stroke risk factors. Peripheral blood samples from 43 acute ischemic stroke patients and 26 control subjects matched on race and gender were used for flow cytometry and complete blood count analyses. Subsequently, cytokine levels and gene expression were measured in γδT cells. The number of circulating γδT cells was decreased by almost 50% (p = 0.005) in the stroke patients. γδT cell counts did not correlate with lesion volume on magnetic resonance diffusion-weighted imaging or with clinical severity in the stroke patients, but γδT cells showed elevated levels of IL-17A (p = 0.048). Decreased γδT cell counts were also associated with older age (p = 0.004), pre-existing hypertension (p = 0.0005) and prevalent coronary artery disease (p = 0.03), with pre-existing hypertension being the most significant predictor of γδT cell counts in a multivariable analysis. γδT cells in human ischemic stroke are reduced in number and show elevated levels of IL-17A. A major reduction in γδT lymphocytes also occurs in hypertension and may contribute to the development of hypertension-mediated stroke and vascular disease.     

The p38 Pathway Regulates Oxidative Stress Tolerance by Phosphorylation of Mitochondrial Protein IscU

The p38 pathway is an evolutionarily conserved signaling pathway that responds to a variety of stresses. However, the underlying mechanisms are largely unknown. In the present study, we demonstrate that p38b is a major p38 MAPK involved in the regulation of oxidative stress tolerance in addition to p38a and p38c in Drosophila. We further show the importance of MK2 as a p38-activated downstream kinase in resistance to oxidative stresses. Furthermore, we identified the iron-sulfur cluster scaffold protein IscU as a new substrate of MK2 both in Drosophila cells and in mammalian cells. These results imply a new mechanistic connection between the p38 pathway and mitochondria iron-sulfur clusters.     

Dyskerin,tRNA genes,and condensin tether pericentric chromatin to the spindle axis in mitosis

Condensin is enriched in the pericentromere of budding yeast chromosomes where it is constrained to the spindle axis in metaphase. Pericentric condensin contributes to chromatin compaction, resistance to microtubule-based spindle forces, and spindle length and variance regulation. Condensin is clustered along the spindle axis in a heterogeneous fashion. We demonstrate that pericentric enrichment of condensin is mediated by interactions with transfer ribonucleic acid (tRNA) genes and their regulatory factors. This recruitment is important for generating axial tension on the pericentromere and coordinating movement between pericentromeres from different chromosomes. The interaction between condensin and tRNA genes in the pericentromere reveals a feature of yeast centromeres that has profound implications for the function and evolution of mitotic segregation mechanisms.     

Neurocognitive outcomes in Malawian children exposed to malaria during pregnancy: An observational birth cohort study

BackgroundAnnually 125 million pregnancies are at risk of malaria infection. However, the impact of exposure to malaria in pregnancy on neurodevelopment in children is not well understood. We hypothesized that malaria in pregnancy and associated maternal immune activation result in neurodevelopmental delay in exposed offspring.Methods and findingsBetween April 2014 and April 2015, we followed 421 Malawian mother–baby dyads (median [IQR] maternal age: 21 [19, 28] years) who were previously enrolled (median [IQR] gestational age at enrollment: 19.7 [17.9, 22.1] weeks) in a randomized controlled malaria prevention trial with 5 or 6 scheduled assessments of antenatal malaria infection by PCR. Children were evaluated at 12, 18, and/or 24 months of age with cognitive tests previously validated in Malawi: the Malawi Developmental Assessment Tool (MDAT) and the MacArthur–Bates Communicative Development Inventories (MCAB-CDI). We assessed the impact of antenatal malaria (n [%] positive: 240 [57.3]), placental malaria (n [%] positive: 112 [29.6]), and maternal immune activation on neurocognitive development in children. Linear mixed-effects analysis showed that children exposed to antenatal malaria between 33 and 37 weeks gestation had delayed language development across the 2-year follow-up, as measured by MCAB-CDI (adjusted beta estimate [95% CI], −7.53 [−13.04, −2.02], p = 0.008). Maternal immune activation, characterized by increased maternal sTNFRII concentration, between 33 and 37 weeks was associated with lower MCAB-CDI language score (adjusted beta estimate [95% CI], −8.57 [−13.09, −4.06], p < 0.001). Main limitations of this study include a relatively short length of follow-up and a potential for residual confounding that is characteristic of observational studies.ConclusionsThis mother–baby cohort presents evidence of a relationship between malaria in pregnancy and neurodevelopmental delay in offspring. Malaria in pregnancy may be a modifiable risk factor for neurodevelopmental injury independent of birth weight or prematurity. Successful interventions to prevent malaria during pregnancy may reduce the risk of neurocognitive delay in children.

Andrea Weckman and co-workers study associations between children’s neurodevelopmental outcomes and malaria in pregnancy.