Reduction of APOE accounts for neurobehavioral deficits in fetal alcohol spectrum disorders.

A hallmark of fetal alcohol spectrum disorders (FASD) is neurobehavioral deficits that still do not have effective treatment. Here, we present that reduction of Apolipoprotein E (APOE) is critically involved in neurobehavioral deficits in FASD. We show that prenatal alcohol exposure (PAE) changes chromatin accessibility of Apoe locus, and causes reduction of APOE levels in both the brain and peripheral blood in postnatal mice. Of note, postnatal administration of an APOE receptor agonist (APOE-RA) mitigates motor learning deficits and anxiety in those mice. Several molecular and electrophysiological properties essential for learning, which are altered by PAE, are restored by APOE-RA. Our human genome-wide association study further reveals that the interaction of PAE and a single nucleotide polymorphism in the APOE enhancer which chromatin is closed by PAE in mice is associated with lower scores in the delayed matching-to-sample task in children. APOE in the plasma is also reduced in PAE children, and the reduced level is associated with their lower cognitive performance. These findings suggest that controlling the APOE level can serve as an effective treatment for neurobehavioral deficits in FASD.

Circulating immune complexes and G6PD deficiency predict readmissions for blackwater fever and severe anemia in children with severe malaria in Eastern Uganda.

BACKGROUND: Recently, there has been an unexplained increase in the incidence of blackwater fever (BWF) in Eastern Uganda. In this study, we evaluate the association between immune complexes, glucose-6-phosphate dehydrogenase (G6PD) deficiency, and the occurrence and recurrence of BWF in children with severe malaria (SM). METHODS: Between 2014 and 2017, children aged six months to <4 years hospitalized with SM and community children (CC) were recruited at two hospitals in Central and Eastern Uganda. We measured serum circulating immune complexes (cIC) and their relationship to SM complications and post-discharge outcomes and evaluated effect mediation through G6PD deficiency. RESULTS: 557 children with SM and 101 CC were enrolled. The mean age of children was 2.1 years. Children with SM had higher cIC levels than CC, p<0.001. After controlling for age, sex, and site, cIC were associated with severe anemia, jaundice, and BWF (adjusted odds ratio, 95% confidence interval: 7.33 (3.45, 15.58), p<0.0001; 4.31 (1.68, 11.08), p=0.002; and 5.21 (2.06, 13.18), p<0.0001), respectively. cIC predicted readmissions for SM, severe anemia, and BWF (adjusted incidence rate ratios (95% confidence interval): 2.11 (1.33, 3.34), p=0.001; 8.62 (2.80, 26.59), p<0.0001; and 7.66 (2.62, 22.45), p<0.0001), respectively. The relationship was most evident in males where the frequency of the G6PD African allele (A-) was 16.8%. G6PD deficiency was associated with increases in cIC in males (p=0.01) and mediation analysis suggested G6PD deficiency contributes to recurrent severe anemia and BWF via increased cIC. CONCLUSIONS: Immune complexes are associated with hemolytic complications and predict recurrences in SM survivors.

Elevated extracellular inorganic phosphate inhibits ecto-phosphatase activity in breast cancer cells: Regulation by hydrogen peroxide.

For cells to obtain inorganic phosphate, ectoenzymes in the plasma membrane, which contain a catalytic site facing the extracellular environment, hydrolyze phosphorylated molecules. In this study, we show that increased Pi levels in the extracellular environment promote a decrease in ecto-phosphatase activity, which is associated with Pi-induced oxidative stress. High levels of Pi inhibit ecto-phosphatase because Pi generates H2 O2 . Ecto-phosphatase activity is inhibited by H2 O2 , and this inhibition is selective for phospho-tyrosine hydrolysis. Additionally, it is shown that the mechanism of inhibition of ecto-phosphatase activity involves lipid peroxidation. In addition, the inhibition of ecto-phosphatase activity by H2 O2 is irreversible. These findings have new implications for understanding ecto-phosphatase regulation in the tumor microenvironment. H2 O2 stimulated by high Pi inhibits ecto-phosphatase activity to prevent excessive accumulation of extracellular Pi, functioning as a regulatory mechanism of Pi variations in the tumor microenvironment.

Ectonucleotidase activity driven by acid ectophosphatase in luminal A MCF-7 breast cancer cells.

Ectophosphatases catalyse the hydrolysis of phosphorylated molecules, such as phospho-amino acids, in the extracellular environment. Nevertheless, the hydrolysis of nucleotides in the extracellular environment is typically catalysed by ectonucleotidases. Studies have shown that acid ectophosphatase, or transmembrane-prostatic acid phosphatase (TM-PAP), a membrane-bound splice variant of prostatic acid phosphatase, has ecto-5'-nucleotidase activity. Furthermore, it was demonstrated that ectophosphatase cannot hydrolyse ATP, ADP, or AMP in triple-negative breast cancer cells. In contrast to previous findings in MDA-MB-231 cells, the ectophosphatase studied in the present work displayed a remarkable capacity to hydrolyse AMP in luminal A breast cancer cells (MCF-7). We showed that AMP dose-dependently inhibited p-nitrophenylphosphate (p-NPP) hydrolysis. The p-NPP and AMP hydrolysis showed similar biochemical behaviours, such as increased hydrolysis under acidic conditions and comparable inhibition by NiCl2, ammonium molybdate, and sodium orthovanadate. In addition, this ectophosphatase with ectonucleotidase activity was essential for the release of adenosine and inorganic phosphate from phosphorylated molecules available in the extracellular microenvironment. This is the first study to show that prostatic acid phosphatase on the membrane surface of breast cancer cells (MCF-7) is correlated with cell adhesion and migration.

Comparative characterisation of an ecto-5'-nucleotidase (CD73) in non-tumoral MCF10-A breast cells and triple-negative MDA-MB-231 breast cancer cells.

Ecto-5'-nucleotidase (CD73) hydrolyses 5'AMP to adenosine and inorganic phosphate. Breast cancer cells (MDA-MB-231) express high CD73 levels, and this enzyme has been found to play a tumour-promoting role in breast cancer. However, no studies have sought to investigate whether CD73 has differential affinity or substrate preferences between noncancerous and cancerous breast cells. In the present study, we aimed to biochemically characterise ecto-5'-nucleotidase in breast cancer cell lines and assess whether its catalytic function and tumour progression are correlated in breast cancer cells. The results showed that compared to nontumoral breast MCF-10A cells, triple-negative breast cancer MDA-MB-231 cells had a higher ecto-5'-nucleotidase expression level and enzymatic activity. Although ecto-5'-nucleotidase activity in the MDA-MB-231 cell line showed no selectivity among monophosphorylated substrates, 5'AMP was preferred by the MCF-10A cell line. Compared to the MCF-10A cell line, the MDA-MB-231 cell line has better hydrolytic ability, lower substrate affinity, and high inhibitory potential after treatment with a specific CD73 inhibitor α,ß­methylene ADP (APCP). Therefore, we demonstrated that a specific inhibitor of the ecto-5-nucleotidase significantly reduced the migratory and invasive capacity of MDA-MB-231 cells, suggesting that ecto-5-nucleotidase activity might play an important role in metastatic progression.

Pathogenic variants in the Longitudinal Early-onset Alzheimer's Disease Study cohort.

INTRODUCTION: One goal of the Longitudinal Early-onset Alzheimer's Disease Study (LEADS) is to investigate the genetic etiology of early onset (40-64 years) cognitive impairment. Toward this goal, LEADS participants are screened for known pathogenic variants. METHODS: LEADS amyloid-positive early-onset Alzheimer's disease (EOAD) or negative early-onset non-AD (EOnonAD) cases were whole exome sequenced (N = 299). Pathogenic variant frequency in APP, PSEN1, PSEN2, GRN, MAPT, and C9ORF72 was assessed for EOAD and EOnonAD. Gene burden testing was performed in cases compared to similar-age cognitively normal controls in the Parkinson's Progression Markers Initiative (PPMI) study. RESULTS: Previously reported pathogenic variants in the six genes were identified in 1.35% of EOAD (3/223) and 6.58% of EOnonAD (5/76). No genes showed enrichment for carriers of rare functional variants in LEADS cases. DISCUSSION: Results suggest that LEADS is enriched for novel genetic causative variants, as previously reported variants are not observed in most cases. HIGHLIGHTS: Sequencing identified eight cognitively impaired pathogenic variant carriers. Pathogenic variants were identified in PSEN1, GRN, MAPT, and C9ORF72. Rare variants were not enriched in APP, PSEN1/2, GRN, and MAPT. The Longitudinal Early-onset Alzheimer's Disease Study (LEADS) is a key resource for early-onset Alzheimer's genetic research.

Effects of Anodal Transcranial Direct Current Stimulation on Training Volume and Pleasure Responses in the Back Squat Exercise Following a Bench Press.

ABSTRACT: Rodrigues, GM, Machado, S, Faria Vieira, LA, Ramalho de Oliveira, BR, Jesus Abreu, MA, Marquez, G, Maranhão Neto, GA, and Lattari, E. Effects of anodal transcranial direct current stimulation on training volume and pleasure responses in the back squat exercise following a bench press. J Strength Cond Res 36(11): 3048-3055, 2022-This study aimed to investigate the effects of anodal transcranial direct current stimulation (a-tDCS) on volume-load and pleasure responses in a back squat following a bench press. Twelve male subjects advanced in resistance training (RT) (age, 25.5 ± 4.4 years) completed 2 experimental trials in a counterbalanced crossover design: a-tDCS and sham conditions. The stimulus was applied over the left dorsolateral prefrontal cortex for 20 minutes using a 2-mA current intensity in a-tDCS condition and 1 minute of active stimulus in the sham condition. Immediately after stimulation, subjects performed the bench press followed by the back squat. The exercise protocol consisted of 3 sets of maximum repetitions at an intensity of 80% of 1 repetition maximum. The volume-load, perceived pleasure, and arousal responses were measured during the RT protocol. The results indicated that volume-load was higher in the a-tDCS condition than in the sham condition for both exercises ( p = 0.02), with large effect for the back squat ( p = 0.045; d = 0.96). The higher volume-load was obtained by increasing the number of repetitions across all sets for the bench press ( p ≤ 0.0001) and only in the first set for the back squat ( p = 0.01). The circumplex model analysis showed a higher pleasure in the bench press and a tendency toward a higher pleasure in the a-tDCS condition. Anodal tDCS may be used as an ergogenic resource for increasing the back squat volume after performing the bench press in resistance-trained male subjects.

Resveratrol is an inhibitor of sodium-dependent inorganic phosphate transport in triple-negative MDA-MB-231 breast cancer cells.

Metastasis is a major cause of death in patients with breast cancer. A growing body of evidence has demonstrated the antitumour effects of resveratrol, a non-flavonoid polyphenol. Resveratrol inhibits metastatic processes, such as the migration and invasion of cancer cells. In several cancer types, the importance of inorganic phosphate (Pi) for tumor progression has been demonstrated. The metastatic process in breast cancer is associated with Na+ -dependent Pi transporters. In this study, we demonstrate, for the first time, that resveratrol inhibits the Na+ -dependent Pi transporter. Results from kinetic analysis shows that resveratrol inhibits Na+ -dependent Pi transport non-competitively. Resveratrol also inhibits adhesion/migration in MDA-MB-231 cells, likely related to inhibition of the Na+ -dependent Pi transporter.

Ectophosphatase activity in the triple-negative breast cancer cell line MDA-MB-231.

Breast cancer is one of the most common cancers in the female population worldwide, and its development is thought to be associated with genetic mutations that lead to uncontrolled and accelerated growth of breast cells. This abnormal behavior requires extra energy, and indeed, tumor cells display a rewired energy metabolism compared to normal breast cells. Inorganic phosphate (Pi) is a glycolytic substrate of glyceraldehyde-3-phosphate dehydrogenase and has an important role in cancer cell proliferation. For cells to obtain Pi, ectoenzymes in the plasma membrane with their catalytic site facing the extracellular environment can hydrolyze phosphorylated molecules, and this is an initial and possibly limiting step for the uptake of Pi by carriers that behave as adjuvants in the process of energy harvesting and thus partially contributes to tumor energy requirements. In this study, the activity of an ectophosphatase in MDA-MB-231 cells was biochemically characterized, and the results showed that the activity of this enzyme was higher in the acidic pH range and that the enzyme had a Km = 4.5 ± 0.5 mM para-nitrophenylphosphate and a Vmax = 2280 ± 158 nM × h-1 × mg protein-1 . In addition, classical acid phosphatase inhibitors, including sodium orthovanadate, decreased enzymatic activity. Sodium orthovanadate was able to inhibit ectophosphatase activity while also inhibiting cell proliferation, adhesion, and migration, which are important processes in tumor progression, especially in metastatic breast cancer MDA-MB-231 cells that have higher ectophosphatase activity than MCF-7 and MCF-10 breast cells.

Extracellular Inorganic Phosphate-Induced Release of Reactive Oxygen Species: Roles in Physiological Processes and Disease Development.

Inorganic phosphate (Pi) is an essential nutrient for living organisms and is maintained in equilibrium in the range of 0.8-1.4 mM Pi. Pi is a source of organic constituents for DNA, RNA, and phospholipids and is essential for ATP formation mainly through energy metabolism or cellular signalling modulators. In mitochondria isolated from the brain, liver, and heart, Pi has been shown to induce mitochondrial reactive oxygen species (ROS) release. Therefore, the purpose of this review article was to gather relevant experimental records of the production of Pi-induced reactive species, mainly ROS, to examine their essential roles in physiological processes, such as the development of bone and cartilage and the development of diseases, such as cardiovascular disease, diabetes, muscle atrophy, and male reproductive system impairment. Interestingly, in the presence of different antioxidants or inhibitors of cytoplasmic and mitochondrial Pi transporters, Pi-induced ROS production can be reversed and may be a possible pharmacological target.

Hydrogen Peroxide Generation as an Underlying Response to High Extracellular Inorganic Phosphate (Pi) in Breast Cancer Cells.

According to the growth rate hypothesis (GRH), tumour cells have high inorganic phosphate (Pi) demands due to accelerated proliferation. Compared to healthy individuals, cancer patients present with a nearly 2.5-fold higher Pi serum concentration. In this work, we show that an increasing concentration of Pi had the opposite effect on Pi-transporters only in MDA-MB-231 when compared to other breast cell lines: MCF-7 or MCF10-A (non-tumoural breast cell line). Here, we show for the first time that high extracellular Pi concentration mediates ROS production in TNBC (MDA-MB-231). After a short-time exposure (1 h), Pi hyperpolarizes the mitochondrial membrane, increases mitochondrial ROS generation, impairs oxygen (O2) consumption and increases PKC activity. However, after 24 h Pi-exposure, the source of H2O2 seems to shift from mitochondria to an NADPH oxidase enzyme (NOX), through activation of PKC by H2O2. Exogenous-added H2O2 modulated Pi-transporters the same way as extracellular high Pi, which could be reversed by the addition of the antioxidant N-acetylcysteine (NAC). NAC was also able to abolish Pi-induced Epithelial-mesenchymal transition (EMT), migration and adhesion of MDA-MB-231. We believe that Pi transporters support part of the energy required for the metastatic processes stimulated by Pi and trigger Pi-induced H2O2 production as a signalling response to promote cell migration and adhesion.

The Roles of Sodium-Independent Inorganic Phosphate Transporters in Inorganic Phosphate Homeostasis and in Cancer and Other Diseases.

Inorganic phosphate (Pi) is an essential nutrient for the maintenance of cells. In healthy mammals, extracellular Pi is maintained within a narrow concentration range of 0.70 to 1.55 mM. Mammalian cells depend on Na+/Pi cotransporters for Pi absorption, which have been well studied. However, a new type of sodium-independent Pi transporter has been identified. This transporter assists in the absorption of Pi by intestinal cells and renal proximal tubule cells and in the reabsorption of Pi by osteoclasts and capillaries of the blood-brain barrier (BBB). Hyperphosphatemia is a risk factor for mineral deposition, the development of diseases such as osteoarthritis, and vascular calcifications (VCs). Na+-independent Pi transporters have been identified and biochemically characterized in vascular smooth muscle cells (VSMCs), chondrocytes, and matrix vesicles, and their involvement in mineral deposition in the extracellular microenvironment has been suggested. According to the growth rate hypothesis, cancer cells require more phosphate than healthy cells due to their rapid growth rates. Recently, it was demonstrated that breast cancer cells (MDA-MB-231) respond to high Pi concentration (2 mM) by decreasing Na+-dependent Pi transport activity concomitant with an increase in Na+-independent (H+-dependent) Pi transport. This Pi H+-dependent transport has a fundamental role in the proliferation and migratory capacity of MDA-MB-231 cells. The purpose of this review is to discuss experimental findings regarding Na+-independent inorganic phosphate transporters and summarize their roles in Pi homeostasis, cancers and other diseases, such as osteoarthritis, and in processes such as VC.

Protein complexes in bacteria.

Large-scale analyses of protein complexes have recently become available for Escherichia coli and Mycoplasma pneumoniae, yielding 443 and 116 heteromultimeric soluble protein complexes, respectively. We have coupled the results of these mass spectrometry-characterized protein complexes with the 285 "gold standard" protein complexes identified by EcoCyc. A comparison with databases of gene orthology, conservation, and essentiality identified proteins conserved or lost in complexes of other species. For instance, of 285 "gold standard" protein complexes in E. coli, less than 10% are fully conserved among a set of 7 distantly-related bacterial "model" species. Complex conservation follows one of three models: well-conserved complexes, complexes with a conserved core, and complexes with partial conservation but no conserved core. Expanding the comparison to 894 distinct bacterial genomes illustrates fractional conservation and the limits of co-conservation among components of protein complexes: just 14 out of 285 model protein complexes are perfectly conserved across 95% of the genomes used, yet we predict more than 180 may be partially conserved across at least half of the genomes. No clear relationship between gene essentiality and protein complex conservation is observed, as even poorly conserved complexes contain a significant number of essential proteins. Finally, we identify 183 complexes containing well-conserved components and uncharacterized proteins which will be interesting targets for future experimental studies.

Genome-wide meta-analyses of cross substance use disorders in European, African, and Latino ancestry populations.

Genetic risks for substance use disorders (SUDs) are due to both SUD-specific and SUD-shared genes. We performed the largest multivariate analyses to date to search for SUD-shared genes using samples of European (EA), African (AA), and Latino (LA) ancestries. By focusing on variants having cross-SUD and cross-ancestry concordant effects, we identified 45 loci. Through gene-based analyses, gene mapping, and gene prioritization, we identified 250 SUD-shared genes. These genes are highly expressed in amygdala, cortex, hippocampus, hypothalamus, and thalamus, primarily in neuronal cells. Cross-SUD concordant variants explained ~ 50% of the heritability of each SUD in EA. The top 5% individuals having the highest polygenic scores were approximately twice as likely to have SUDs as others in EA and LA. Polygenic scores had higher predictability in females than in males in EA. Using real-world data, we identified five drugs targeting identified SUD-shared genes that may be repurposed to treat SUDs.

Inorganic Phosphate (Pi) in the Breast Cancer Microenvironment: Production, Transport and Signal Transduction as Potential Targets for Anticancer Strategies.

Tumor cells develop a high demand for inorganic phosphate (Pi) due to their high growth rates and energy requirements. Serum Pi concentrations in cancer patients have been found to be two to four times higher than baseline levels in healthy individuals. Twofold Pi accumulation was observed in breast cancer cells in the mouse tumor microenvironment. In the breast tumoral microenvironment, ectonucleotidases and ectophosphatases-presenting catalytic sites facing the extracellular environment-could be involved in the extracellular release of Pi to be internalized by Pi transporters to fuel the high energy requirement typical of cancer cells. Two Pi transporters were characterized in breast cancer cells (Na+-dependent and H+-dependent) with strong associations with tumor processes such as proliferation, migration, adhesion, and epithelium-mesenchymal transition (EMT). Moreover, a high extracellular Pi concentration stimulates ROS production in triple-negative breast cancer cells by Pi transport stimulation. Several compounds show a potent ability to inhibit ectonucleotidases, ectophosphatases, Pi transporters, and Pi-modulated signal pathways in breast cancer cells and regulate proliferation, migration, adhesion, and EMT. This review article aimed to gather the relevant experimental records regarding Pi's effects on the breast cancer microenvironment and points to possible inhibitors for ectonucleotidases, ectophosphatases, Pi transporters, and Pi-modulated signal pathways as potential chemotherapeutic agents or Pi acting as a potent enhancer of classical chemical-induced cytotoxicity in triple-negative breast cancer cells.

Anodal Transcranial Direct Current Stimulation Does Not Affect Velocity Loss During a Typical Resistance Exercise Session.

Purpose: This study investigated the effects of transcranial direct current stimulation (tDCS) on velocity loss in a typical resistance exercise session. Methods: Twelve recreationally resistance-trained males (age = 24.8 ± 3.0 years, body mass = 78.9 ± 13.6 kg, and height = 174.3 ± 7.3 cm) completed two experimental trials in a counterbalanced crossover design: anodal tDCS and sham conditions. The stimuli were applied over the left dorsolateral prefrontal cortex for 20 minutes, using a 2 mA current intensity in anodal tDCS and a 1-minute active stimulus in the sham condition. After stimulation, subjects performed three sets of the bench press at a 70% of 1 maximum repetition intensity and 1 min of inter-set rest. The velocity loss was calculated as the relative difference between the fastest repetition velocity (usually first) and the velocity of the last repetition of each set and averaged over all three sets. Results: The results found no interaction between conditions and sets (P = .122), and no effect for conditions (P = .323) or sets (P = .364) for the velocity loss in each set. Also, no differences were found between the average velocity loss of the three sets in the anodal tDCS (-25.0 ± 4.7%) and sham condition (-23.3 ± 6.4%; P = .323). Conclusion: Anodal tDCS does not affect movement velocity in a typical strength training protocol in recreationally trained subjects.

Mutation in the Bone Morphogenetic Protein signaling pathway sensitize zebrafish and humans to ethanol-induced jaw malformations.

Fetal Alcohol Spectrum Disorders (FASD) describe ethanol-induced developmental defects including craniofacial malformations. While ethanol-sensitive genetic mutations contribute to facial malformations, the impacted cellular mechanisms remain unknown. Bmp signaling is a key regulator of epithelial morphogenesis driving facial development, providing a possible ethanol-sensitive mechanism. We found that zebrafish mutants for Bmp signaling components are ethanol-sensitive and affect anterior pharyngeal endoderm shape and gene expression, indicating ethanol-induced malformations of the anterior pharyngeal endoderm cause facial malformations. Integrating FASD patient data, we provide the first evidence that variants in the human Bmp receptor gene BMPR1B associate with ethanol-related differences in jaw volume. Our results show that ethanol exposure disrupts proper morphogenesis of, and tissue interactions between, facial epithelia that mirror overall viscerocranial shape changes and are predictive for Bmp-ethanol associations in human jaw development. Our data provide a mechanistic paradigm linking ethanol to disrupted epithelial cell behaviors that underlie facial defects in FASD.

Transcranial Direct Current Stimulation Combined With or Without Caffeine: Effects on Training Volume and Pain Perception.

Purpose: This study aimed to investigate the acute effects of tDCS combined with caffeine intake on training volume and pain perception in the bench press in resistance-trained males. The correlation between training volume and pain perception was also assessed in all interventions. Methods: Sixteen healthy males (age = 25.2 ± 4.7 years, body mass = 82.8 ± 9.1 kg, and height = 178.3 ± 5.7 cm), advanced in RT, were randomized and counterbalanced for the following experimental conditions: Sham tDCS with placebo intake (Sham+Pla), Sham tDCS with caffeine intake (Sham+Caff), anodal tDCS with placebo intake (a-tDCS+Pla), and anodal tDCS with caffeine intake (a-tDCS+Caff). The caffeine or placebo ingestion (both with 5 mg.kg-1) occurred 40 minutes before the tDCS sessions. The tDCS was applied over the left DLPFC for 20 minutes, with a 2 mA current intensity. After the tDCS sessions, participants performed the bench press with an 80% of 1RM load, where training volume and pain perception were measured. Results: Training volume was higher in the 1st and 2nd sets in both a-tDCS+Caff and Sham+Caff conditions, compared to the Sham+Pla condition (P < .05). Both a-tDCS+Caff and a-tDCS+Pla showed an increased pain perception during the third set compared to the first set. Also, no correlation was found between the number of repetitions and pain perception in any condition (P > .05). Conclusion: This research revealed that caffeine intake alone could be used as an ergogenic aid during resistance training programs in resistance-trained males.

Functional characterization of maternally accumulated hydrolases in the mature oocytes of the vector Rhodnius prolixus reveals a new protein phosphatase essential for the activation of the yolk mobilization and embryo development.

Yolk biogenesis and consumption have been well conserved in oviparous animals throughout evolution. Most egg-laying animals store yolk proteins within the oocytes' yolk granules (Ygs). Following fertilization, the Ygs participate in controlled pathways of yolk breakdown to support the developing embryo's anabolic metabolism. While the unfolding of the yolk degradation program is a crucial process for successful development in many species, the molecular mechanisms responsible for yolk mobilization are still mysterious and have mostly not been explored. Here, we investigate the functional role of the oocyte maternally accumulated mRNAs of a protein phosphatase (PP501) and two aspartic proteases (cathepsin-D 405, CD405 and cathepsin-D 352, CD352) in the yolk degradation and reproduction of the insect vector of Chagas disease Rhodnius prolixus. We found that PP501 and CD352 are highly expressed in the vitellogenic ovary when compared to the other organs of the adult insect. Parental RNAi silencing of PP501 resulted in a drastic reduction in oviposition and increased embryo lethality whereas the silencing of CD352 resulted only in a slight decrease in oviposition and embryo viability. To further investigate the PP501-caused high reproduction impairment, we investigated the Ygs biogenesis during oocyte maturation and the activation of the yolk degradation program at early development. We found that the Ygs biogenesis was deficient during oogenesis, as seen by flow cytometry, and that, although the PP501-silenced unviable eggs were fertilized, the Ygs acidification and acid phosphatase activity were affected, culminating in a full impairment of the yolk proteins degradation at early embryogenesis. Altogether we found that PP501 is required for the oocyte maturation and the activation of the yolk degradation, being, therefore, essential for this vector reproduction.

Iron Uptake Controls Trypanosoma cruzi Metabolic Shift and Cell Proliferation.

(1) Background: Ionic transport in Trypanosoma cruzi is the object of intense studies. T. cruzi expresses a Fe-reductase (TcFR) and a Fe transporter (TcIT). We investigated the effect of Fe depletion and Fe supplementation on different structures and functions of T. cruzi epimastigotes in culture. (2) Methods: We investigated growth and metacyclogenesis, variations of intracellular Fe, endocytosis of transferrin, hemoglobin, and albumin by cell cytometry, structural changes of organelles by transmission electron microscopy, O2 consumption by oximetry, mitochondrial membrane potential measuring JC-1 fluorescence at different wavelengths, intracellular ATP by bioluminescence, succinate-cytochrome c oxidoreductase following reduction of ferricytochrome c, production of H2O2 following oxidation of the Amplex® red probe, superoxide dismutase (SOD) activity following the reduction of nitroblue tetrazolium, expression of SOD, elements of the protein kinase A (PKA) signaling, TcFR and TcIT by quantitative PCR, PKA activity by luminescence, glyceraldehyde-3-phosphate dehydrogenase abundance and activity by Western blotting and NAD+ reduction, and glucokinase activity recording NADP+ reduction. (3) Results: Fe depletion increased oxidative stress, inhibited mitochondrial function and ATP formation, increased lipid accumulation in the reservosomes, and inhibited differentiation toward trypomastigotes, with the simultaneous metabolic shift from respiration to glycolysis. (4) Conclusion: The processes modulated for ionic Fe provide energy for the T. cruzi life cycle and the propagation of Chagas disease.