A dual-function SNF2 protein drives chromatid resolution and nascent transcripts removal in mitosis.

Mitotic chromatin is largely assumed incompatible with transcription due to changes in the transcription machinery and chromosome architecture. However, the mechanisms of mitotic transcriptional inactivation and their interplay with chromosome assembly remain largely unknown. By monitoring ongoing transcription in Drosophila early embryos, we reveal that eviction of nascent mRNAs from mitotic chromatin occurs after substantial chromosome compaction and is not promoted by condensin I. Instead, we show that the timely removal of transcripts from mitotic chromatin is driven by the SNF2 helicase-like protein Lodestar (Lds), identified here as a modulator of sister chromatid cohesion defects. In addition to the eviction of nascent transcripts, we uncover that Lds cooperates with Topoisomerase 2 to ensure efficient sister chromatid resolution and mitotic fidelity. We conclude that the removal of nascent transcripts upon mitotic entry is not a passive consequence of cell cycle progression and/or chromosome compaction but occurs via dedicated mechanisms with functional parallelisms to sister chromatid resolution.

Histone Deacetylase Inhibitors Protect Against Pyruvate Dehydrogenase Dysfunction in Huntington's Disease.

Transcriptional deregulation and changes in mitochondrial bioenergetics, including pyruvate dehydrogenase (PDH) dysfunction, have been described in Huntington's disease (HD). We showed previously that the histone deacetylase inhibitors (HDACIs) trichostatin A and sodium butyrate (SB) ameliorate mitochondrial function in cells expressing mutant huntingtin. In this work, we investigated the effect of HDACIs on the regulation of PDH activity in striatal cells derived from HD knock-in mice and YAC128 mice. Mutant cells exhibited decreased PDH activity and increased PDH E1alpha phosphorylation/inactivation, accompanied by enhanced protein levels of PDH kinases 1 and 3 (PDK1 and PDK3). Exposure to dichloroacetate, an inhibitor of PDKs, increased mitochondrial respiration and decreased production of reactive oxygen species in mutant cells, emphasizing PDH as an interesting therapeutic target in HD. Treatment with SB and sodium phenylbutyrate, another HDACI, recovered cell viability and overall mitochondrial metabolism in mutant cells. Exposure to SB also suppressed hypoxia-inducible factor-1 (HIF-1α) stabilization and decreased the transcription of the two most abundant PDK isoforms, PDK2 and PDK3, culminating in increased PDH activation in mutant cells. Concordantly, PDK3 knockdown improved mitochondrial function, emphasizing the role of PDK3 inactivation on the positive effects achieved by SB treatment. YAC128 mouse brain presented higher mRNA levels of PDK1-3 and PDH phosphorylation and decreased energy levels that were significantly ameliorated after SB treatment. Furthermore, enhanced motor learning and coordination were observed in SB-treated YAC128 mice. These results suggest that HDACIs, particularly SB, promote the activity of PDH in the HD brain, helping to counteract HD-related deficits in mitochondrial bioenergetics and motor function.SIGNIFICANCE STATEMENT The present work provides a better understanding of mitochondrial dysfunction in Huntington's disease (HD) by showing that the pyruvate dehydrogenase (PDH) complex is a promising therapeutic target. In particular, the histone deacetylase inhibitor sodium butyrate (SB) may indirectly (through reduced hypoxia-inducible factor 1 alpha stabilization) decrease the expression of the most abundant PDH kinase isoforms (e.g., PDK3), ameliorating PDH activity and mitochondrial metabolism and further affecting motor behavior in HD mice, thus constituting a promising agent for HD neuroprotective treatment.

Mitochondrial Dysfunction in Huntington's Disease.

Mitochondrial dysfunction has been described as an early pathological mechanism delineating the selective neurodegeneration that occurs in Huntington's disease (HD), a polyglutamine-expansion disorder that largely affects the striatum and the cerebral cortex. Over the years, mitochondria roles in eukaryotic cells (e.g. in neurons) have largely diverged from the classically attributed cell power source; indeed, mitochondria not only contribute for synthesis of several metabolites, but are also dynamic organelles that fragment and fuse to achieve a maximal bioenergetic performance, are transported along microtubules, regulate intracellular calcium homeostasis through the interaction with the endoplasmic reticulum, produce free radicals and participate in cell death processes. Indeed, most of these activities have been demonstrated to be affected in HD, potentially contributing for the neuronal dysfunction in pre-symptomatic stages. This chapter resumes some of the evidences that pose mitochondria as a main regulatory organelle in HD-affected neurons, uncovering some potentially therapeutic mitochondrial-based relevant targets.

Carbon Threads Supercapacitors for Washable e-Textile Applications: Configurations and Electrochemical Performance.

Technological solutions for emerging e-textiles are being sought to enable e-wear technology to be self-sustaining and lightweight. A rippling 1D carbon fiber capacitor design was made with commercial carbon threads as electrodes using simulated sweat solution as the electrolyte. This is particularly relevant for potential sports textile applications in which sweat could serve as an electrochemical energy source. An electrospun cellulose acetate fiber membrane and a commercially available felt were used as separators capable of soaking the electrolyte. These were tested in braided and woven electrode configurations, respectively. Functionalizing the carbon wires with polypyrrole (PPy) enhanced the surface area and significantly increased the specific capacity by approximately an order of magnitude (0.62 F/g). Cyclic voltammetry and charge-discharge tests confirmed the washability and durability of the devices for at least 1000 cycles.

How Do Healthy Women Perceive the Risk of Breast Cancer? The Role of Illness Perceptions and Compared Risk between Portugal and the U.A.E.

(1) Background: Breast cancer (BC) shows significant epidemiological differences between Eastern and Western countries. These may arise from socio-cultural factors influencing how healthy young women perceive this condition, their risk of getting cancer, and the implications for preventive screening behaviors. In this study, the illness perceptions, individual risk perception, compared risk, and beliefs about preventive behaviors for BC of female university students were compared using an anonymous online survey between a European country (Portugal) and the United Arab Emirates. (2) Method: A structural equation model (SEM) was developed to investigate the hypothetical relationship between illness perceptions and compared risk as predictors of perceived risk for BC. (3) Results: There were significant differences between the study variables. The SEM was invariant, but the differences between regression coefficients in both countries were highly statistically significant. Mediation analyses revealed a significant indirect effect of compared risk on individual risk and a significantly stronger direct effect for the Emirati sample. (4) Conclusions: These findings suggest that cultural research may help to explain factors that may shape social comparison of individual risk characteristics and influence perceived risk. Moreover, providing culturally appropriate strategies to be designed and implemented can promote early detection behaviors for BC.

Immediate Effects of Hypnosis, Mindfulness Meditation, and Prayer on Cold Pressor Outcomes: A Four-Arm Parallel Experimental Study.

Purpose: Previous research supports the usefulness of hypnosis (HYP), mindfulness meditation (MM), and prayer as pain self-management strategies in adults with chronic pain. However, their effects on acute pain have been less researched, and no previous head-to-head study compared the immediate effects of these three approaches on pain-related outcomes. This study compared the immediate effects of HYP, MM, and Christian prayer (CP) on pain intensity, pain tolerance, and stress as assessed by heart rate variability (HRV). Participants and Methods: A total of 232 healthy adults were randomly assigned to, and completed, a single 20-minute session of MM, SH, CP, or an attention control (CN), and underwent two cycles (one pre- and one post-intervention) of Cold Pressor Arm Wrap (CPAW). Sessions were audio-delivered. Participants responded to pre- and post-intervention pain intensity measurements. Pain tolerance (sec) was assessed during the CPAW cycles. HRV was assessed at baseline, and at pre- and post-intervention CPAW cycles. The study protocol was pre-registered at the ClinicalTrials.gov registry (NCT04491630). Results: Small within-group decreases in pain intensity and small increases in pain tolerance were found for HYP and MM from the pre- to the post-intervention. Small within-group improvements in the LH/HF ratio were also found for HYP. The exploratory between-group pairwise comparisons revealed a medium effect size effects of HYP on pain tolerance relative to the control condition. The effects of CP were positive, but small and not statistically significant. Only small to medium, though non-significant, Time × Group interaction effects were found. Conclusion: Study results suggest that single short-term HYP and MM sessions, but not biblical-based CP, may be useful for acute pain self-management, with HYP being the slightly superior option. Future research should compare the effects of different types of prayer and examine the predictors and moderators of these pain approaches' effects on pain-related outcomes.

Mitochondrial SIRT3 confers neuroprotection in Huntington's disease by regulation of oxidative challenges and mitochondrial dynamics.

SIRT3 is a major regulator of mitochondrial acetylome. Here we show that SIRT3 is neuroprotective in Huntington's disease (HD), a motor neurodegenerative disorder caused by an abnormal expansion of polyglutamines in the huntingtin protein (HTT). Protein and enzymatic analysis revealed that increased SIRT3 is a signature in several HD models, including human HD brain, which is regulated by oxidative species. While loss of SIRT3 further aggravated the oxidative phenotype, antioxidant treatment regularized SIRT3 levels. SIRT3 overexpression promoted the antioxidant effect in cells expressing mutant HTT, leading to enhanced mitochondrial function and balanced dynamics. Decreased Fis1 and Drp1 accumulation in mitochondria induced by SIRT3 expression favored mitochondrial elongation, while the SIRT3 activator ε-viniferin improved anterograde mitochondrial neurite transport, sustaining cell survival. Notably, SIRT3 fly-ortholog dSirt2 overexpression in HD flies ameliorated neurodegeneration and extended lifespan. These findings provide a link between oxidative stress and mitochondrial dysfunction hypotheses in HD and offer an opportunity for therapeutic development.

Microinjection Techniques in Fly Embryos to Study the Function and Dynamics of SMC Complexes.

Structural maintenance of chromosomes (SMC) proteins are critical to maintain mitotic fidelity in all organisms. Over the last decades, acute inactivation of these complexes, together with the analysis of their dynamic binding to mitotic chromatin, has provided important insights on the molecular mechanism of these complexes as well as into the consequences of their failure at different stages of mitosis.Here, we describe a methodology to study both SMC function and dynamics using Drosophila melanogaster syncytial embryos. This system presents several advantages over canonical inactivation or imaging approaches. Efficient and fast inactivation of SMC complexes can be achieved by the use of tobacco etch virus (TEV) protease in vivo to cleave engineered versions of the SMC complexes. In contrast to genetically encoded TEV protease expression, Drosophila embryos enable prompt delivery of the protease by microinjection techniques, as detailed here, thereby allowing inactivation of the complexes within few minutes. Such an acute inactivation approach, when coupled with real-time imaging, allows for the analysis of the immediate consequences upon protein inactivation. As described here, this system also presents unique advantages to follow the kinetics of the loading of SMC complexes onto mitotic chromatin. We describe the use of Drosophila embryos to study localization and turnover of these molecules through live imaging and fluorescence recovery after photobleaching (FRAP) approaches.

Assessing Mitochondrial Function in In Vitro and Ex Vivo Models of Huntington's Disease.

Mitochondrial dysfunction has gained a preponderant role in the pathogenesis of Huntington's disease (HD). Mutant huntingtin (mHTT) directly interacts with mitochondria in a deleterious manner. As the central hub of the cell, not only mitochondrial bioenergetics is affected but there is also diminished mitochondrial membrane potential (Δψ m) and altered production of reactive oxygen species (ROS). Restoration of mitochondrial function has proven to be a major player in the search and establishment of therapeutics for HD patients. As such, performing an overall study of mitochondrial function is crucial. In this chapter, we describe some methodologies used to study mitochondrial function by determining the oxygen consumption, changes in Δψ m, mitochondrial calcium handling, and levels of mitochondrial ROS. Here we focus on biological samples derived from HD versus control cells and/or animal models, namely functional isolated brain mitochondria, an ex vivo animal model, and cultured cells, including cell lines and primary neural cultures, as in vitro models.

Comparative Mitochondrial-Based Protective Effects of Resveratrol and Nicotinamide in Huntington's Disease Models.

Sirtuin 1 (SIRT1) is a nicotinamide adenine dinucleotide (NAD+)-dependent lysine deacetylase that regulates longevity and enhances mitochondrial metabolism. Both activation and inhibition of SIRT1 were previously shown to ameliorate neuropathological mechanisms in Huntington's disease (HD), a neurodegenerative disease that selectively affects the striatum and cortex and is commonly linked to mitochondrial dysfunction. Thus, in this study, we tested the influence of resveratrol (RESV, a SIRT1 activator) versus nicotinamide (NAM, a SIRT1 inhibitor) in counteracting mitochondrial dysfunction in HD models, namely striatal and cortical neurons isolated from YAC128 transgenic mice embryos, HD human lymphoblasts, and an in vivo HD model. HD cell models displayed a deregulation in mitochondrial membrane potential and respiration, implicating a decline in mitochondrial function. Further studies revealed decreased PGC-1α and TFAM protein levels, linked to mitochondrial DNA loss in HD lymphoblasts. Remarkably, RESV completely restored these parameters, while NAM increased NAD+ levels, providing a positive add on mitochondrial function in in vitro HD models. In general, RESV decreased while NAM increased H3 acetylation at lysine 9. In agreement with in vitro data, continuous RESV treatment for 28 days significantly improved motor coordination and learning and enhanced expression of mitochondrial-encoded electron transport chain genes in YAC128 mice. In contrast, high concentrations of NAM blocked mitochondrial-related transcription, worsening motor phenotype. Overall, data indicate that activation of deacetylase activity by RESV improved gene transcription associated to mitochondrial function in HD, which may partially control HD-related motor disturbances.

Dynamics of Wolbachia pipientis Gene Expression Across the Drosophila melanogaster Life Cycle.

Symbiotic interactions between microbes and their multicellular hosts have manifold biological consequences. To better understand how bacteria maintain symbiotic associations with animal hosts, we analyzed genome-wide gene expression for the endosymbiotic α-proteobacteria Wolbachia pipientis across the entire life cycle of Drosophila melanogaster. We found that the majority of Wolbachia genes are expressed stably across the D. melanogaster life cycle, but that 7.8% of Wolbachia genes exhibit robust stage- or sex-specific expression differences when studied in the whole-organism context. Differentially-expressed Wolbachia genes are typically up-regulated after Drosophila embryogenesis and include many bacterial membrane, secretion system, and ankyrin repeat-containing proteins. Sex-biased genes are often organized as small operons of uncharacterized genes and are mainly up-regulated in adult Drosophila males in an age-dependent manner. We also systematically investigated expression levels of previously-reported candidate genes thought to be involved in host-microbe interaction, including those in the WO-A and WO-B prophages and in the Octomom region, which has been implicated in regulating bacterial titer and pathogenicity. Our work provides comprehensive insight into the developmental dynamics of gene expression for a widespread endosymbiont in its natural host context, and shows that public gene expression data harbor rich resources to probe the functional basis of the Wolbachia-Drosophila symbiosis and annotate the transcriptional outputs of the Wolbachia genome.

A novel requirement for Janus kinases as mediators of drug resistance induced by fibroblast growth factor-2 in human cancer cells.

The development of resistance to chemotherapy is a major cause of cancer-related death. Elucidating the mechanisms of drug resistance should thus lead to novel therapeutic strategies. Fibroblast growth factor (FGF)-2 signaling induces the assembly of a multi-protein complex that provides tumor cells with the molecular machinery necessary for drug resistance. This complex, which involves protein kinase C (PKC) ε, v-raf murine sarcoma viral oncogene homolog B1 (B-RAF) and p70 S6 kinase ß (S6K2), enhances the selective translation of anti-apoptotic proteins such as B-cell leukaemia/lymphoma-2 (BCL-2) and inhibitors of apoptosis protein (IAP) family members and these are able to protect multiple cancer cell types from chemotherapy-induced cell death. The Janus kinases (JAKs) are most noted for their critical roles in mediating cytokine signaling and immune responses. Here, we show that JAKs have novel functions that support their consideration as new targets in therapies aimed at reducing drug resistance. As an example, we show that the Janus kinase TYK2 is phosphorylated downstream of FGF-2 signaling and required for the full phosphorylation of extracellular signal-regulated kinase (ERK) 1/2. Moreover, TYK2 is necessary for the induction of key anti-apoptotic proteins, such as BCL-2 and myeloid cell leukemia sequence (MCL) 1, and for the promotion of cell survival upon FGF-2. Silencing JAK1, JAK2 or TYK2 using RNA interference (RNAi) inhibits FGF2-mediated proliferation and results in the sensitization of tumor cells to chemotherapy-induced killing. These effects are independent of activation of signal transducer and activator of transcription (STAT) 1, STAT3 and STAT5A/B, the normal targets of JAK signaling. Instead, TYK2 associates with the other kinases previously implicated in FGF-2-mediated drug resistance. In light of these findings we hypothesize that TYK2 and other JAKs are important modulators of FGF-2-driven cell survival and that inhibitors of these kinases will likely improve the effectiveness of other cancer therapies.