Age-Dependent Changes in Transcription Factor FOXO Targeting in Female Drosophila

FOXO transcription factors have long been associated with longevity control and tissue homeostasis. Although the transcriptional regulation of FOXO have been previously characterized (especially in long-lived insulin mutants and under stress conditions), how normal aging impacts the transcriptional activity of FOXO is poorly understood. Here, we conducted a chromatin immunoprecipitation sequencing (ChIP-Seq) analysis in both young (2-week-old) and aged (5-week-old) wild-type female fruit flies, Drosophila melanogaster, to evaluate the dynamics of FOXO gene targeting during aging. Intriguingly, the number of FOXO-bound genes dramatically decreases with age (from 2617 to 224). Consistent to the reduction of FOXO binding activity, many genes targeted by FOXO in young flies are transcriptionally altered with age, either up-regulated (FOXOrepressing genes) or down-regulated (FOXO-activating genes) in adult head tissue. In addition, we show that many FOXO-bound genes in wild-type flies are unique from those in insulin receptor substrate chico mutants. Distinct from chico mutants, FOXO targets specific cellular processes (e.g., actin cytoskeleton) and signaling pathways (e.g., Hippo, MAPK) in young wild-type female flies. FOXO targeting on these pathways decreases with age. Interestingly, FOXO targets in aged flies are enriched in cellular processes like chromatin organization and nucleosome assembly. Furthermore, FOXO binding to core histone genes is well maintained at aged flies. Together, our findings provide new insights into dynamic FOXO targeting under normal aging and highlight the diverse and understudied regulatory mechanisms for FOXO transcriptional activity

Fluorescence detection of cervical intraepithelial neoplasia for photodynamic therapy with the topical agents 5-aminolevulinic acid and benzoporphyrin-derivative monoacid ring

ObjectiveThe aim of this study was to determine whether 2 photosensitizers, benzoporphyrin-derivative monoacid ring and 5-aminolevulinic acid, are selectively absorbed by dysplastic cervical cells after topical administration.Study designThis phase I clinical trial involved 18 women with biopsy-proven cervical intraepithelial neoplasia at the Beckman Laser Institute, Irvine, Calif. Colposcopically directed cervical biopsy specimens obtained after 1.5, 3, or 6 hours of exposure to a randomly assigned photosensitizer were evaluated for selective drug absorption with hematoxylin and eosin staining and fluorescence microscopy.ResultsAfter exposure to 5-aminolevulinic acid, cervical tissue showed maximal fluorescence in dysplastic cells relative to normal cells, with negligible stromal fluorescence. According to our detection methods benzoporphyrin-derivative monoacid ring demonstrated nonselective, diffusion-driven uptake, with fluorescence appearing in the superficial cells, followed by nonselective drug absorption in the remaining cells and stroma of the epithelium.ConclusionOur data demonstrated selective absorption of 5-aminolevulinic acid by dysplastic cervical cells. This agent therefore represents a promising photosensitizing prodrug for the treatment of cervical intraepithelial neoplasia with photodynamic therapy

Prenatal Cocaine Exposure Increases Synaptic Localization of a Neuronal RasGEF, GRASP-1 via Hyperphosphorylation of AMPAR Anchoring Protein, GRIP

Prenatal cocaine exposure causes sustained phosphorylation of the synaptic anchoring protein, glutamate receptor interacting protein (GRIP1/2), preventing synaptic targeting of the GluR2/3-containing alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type glutamate receptors (AMPARs; J. Neurosci. 29: 6308–6319, 2009). Because overexpression of GRIP-associated neuronal rasGEF protein (GRASP-1) specifically reduces the synaptic targeting of AMPARs, we hypothesized that prenatal cocaine exposure enhances GRASP-1 synaptic membrane localization leading to hyper-activation of ras family proteins and heightened actin polymerization. Our results show a markedly increased GRIP1-associated GRASP-1 content with approximately 40% reduction in its rasGEF activity in frontal cortices (FCX) of 21-day-old (P21) prenatal cocaine-exposed rats. This cocaine effect is the result of a persistent protein kinase C (PKC)- and downstream Src tyrosine kinase-mediated GRIP phosphorylation. The hyperactivated PKC also increased membrane-associated GRASP-1 and activated small G-proteins RhoA, cdc42/Rac1 and Rap1 as well as filamentous actin (F-actin) levels without an effect on the phosphorylation state of actin. Since increased F-actin facilitates protein transport, our results suggest that increased GRASP-1 synaptic localization in prenatal cocaine-exposed brains is an adaptive response to restoring the synaptic expression of AMPA-GluR2/3. Our earlier data demonstrated that persistent PKC-mediated GRIP phosphorylation reduces GluR2/3 synaptic targeting in prenatal cocaine-exposed brains, we now show that the increased GRIP-associated GRASP-1 may contribute to the reduction in GluR2/3 synaptic expression and AMPAR signaling defects

Machine learning uncovers the most robust self-report predictors of relationship quality across 43 longitudinal couples studies

Given the powerful implications of relationship quality for health and well-being, a central mission of relationship science is explaining why some romantic relationships thrive more than others. This large-scale project used machine learning (i.e., Random Forests) to 1) quantify the extent to which relationship quality is predictable and 2) identify which constructs reliably predict relationship quality. Across 43 dyadic longitudinal datasets from 29 laboratories, the top relationship-specific predictors of relationship quality were perceived-partner commitment, appreciation, sexual satisfaction, perceived-partner satisfaction, and conflict. The top individual-difference predictors were life satisfaction, negative affect, depression, attachment avoidance, and attachment anxiety. Overall, relationship-specific variables predicted up to 45% of variance at baseline, and up to 18% of variance at the end of each study. Individual differences also performed well (21% and 12%, respectively). Actor-reported variables (i.e., own relationship-specific and individual-difference variables) predicted two to four times more variance than partner-reported variables (i.e., the partner’s ratings on those variables). Importantly, individual differences and partner reports had no predictive effects beyond actor-reported relationship-specific variables alone. These findings imply that the sum of all individual differences and partner experiences exert their influence on relationship quality via a person’s own relationship-specific experiences, and effects due to moderation by individual differences and moderation by partner-reports may be quite small. Finally, relationship-quality change (i.e., increases or decreases in relationship quality over the course of a study) was largely unpredictable from any combination of self-report variables. This collective effort should guide future models of relationships

Changing balances in Dutch higher education

Like many other higher education systems in the Western world, Dutch higher education underwent profound changes during the last decade. In this article we will present an overview of these changes, and try to formulate an analytical framework that might be suited to analyze this process. In order to set the stage, we will begin with an overview of the Dutch higher education system, in which the broad structure is described, and some trends are presented. Next, an overview is given of the retrenchment and restructuring operations with which Dutch higher education was confronted during the last decade. Drawing, mainly, on public administration and political theory, we then attempt to formulate a framework for analysis. In this we focus on the Dutch higher education system as a policy network, and address the relationships that exist between the various key actors in the network: between government and higher education, among higher education institutions themselves, and among the different actors within the institutions, especially administrators and academics. In doing so, we hope to demonstrate that at all these levels some identical basic processes operate which to a large extent determine the outcomes of governmental policies aimed at changing the higher education system. Time and again the modern state stumbles over the academic system (Clark 1983: 137

Analysis of transcription factor FOXO in the regulation of stress, aging, and neuromuscular tissue homeostasis in Drosophila melanogaster

The transcription factor FOXO is a known regulator of tissue homeostasis and animal lifespan. It was first identified in its ability to promote longevity through the insulin signaling pathway, and has since been implicated in numerous cellular processes. Members of the FOXO protein family control a wide array of cellular functions including metabolism, cell cycle arrest, apoptosis, stress resistance, and aging. In response to various signaling cues, FOXO proteins can localize into the nucleus and interact with DNA to regulate transcription. FOXO is known as a longevity gene, however, how FOXO behavior changes during aging is not well understood. Normal aging involves a progressive decline in cell function and an accumulation of oxidative damage. Additionally, with age comes a reduction in FOXO gene expression, and FOXO protein activity can become dysregulated. To resolve how FOXO activity changes with normal aging, we began with chromatin immunoprecipitation sequencing (ChIP-seq) to compare differences in FOXO chromatin binding between young and old organisms, using Drosophila melanogaster as a model. In Drosophila, there is only one homolog representing the FOXO protein family, known as dFOXO. Through our investigation, we found that the number of dFOXO-bound DNA regions decreases with age, and see a number of these targeted genes undergo changes in expression with aging. Some pathways targeted by FOXO at a young age are Hippo, WNT, and MAPK signaling pathways. FOXO is also known to mitigate oxidative stress, which is a contributing factor to age-related cellular degeneration. To understand FOXO dynamics in response to oxidative stress, we used Mass Spectrometry to evaluate dFOXO protein interacting networks under control and stress conditions. We observed a change in several dFOXO partners under paraquat-induced stress such as Stonewall, a chromatin modulator, and Hangover, a key transcription factor regulating neuromuscular junction (NMJ) morphology and neuronal activity. Both FOXO and Hangover show altered NMJ morphologies, and have potentially disrupted vesicle cycling. Given that FOXO has been linked to homeostatic maintenance of neuronal processes across animal species, we performed genetic analysis to investigate how dFOXO regulate NMJ aging in adult flies. Adult NMJs are known to undergo loss of synaptic homeostasis with aging, which causes functional decline and can lead to neurodegeneration. We profiled adult Drosophila abdominal NMJs and found that loss of function FOXO mutants exhibited morphological profiles similar to those of middle aged wild-type flies. We also observed an abnormal accumulation of late endosomes associated with the NMJ both in aged flies and with knockdown of motor neuron FOXO. Overexpression of FOXO can delay this accumulation, suggesting FOXO is a positive regulator of neuronal homeostasis. We performed a genetic screen and identified pathways such as MAPK that act downstream of FOXO to control NMJ homeostasis during aging. Collectively, this thesis provides evidence for global changes in FOXO chromatin binding activity under normal aging, highlights FOXO protein network dynamics that occur under oxidative stress, and displays homeostatic regulation of the adult Drosophila neuromuscular junction. These results illustrate how the transcription factor FOXO acts as a modulator of cellular homeostasis, and how FOXO activity is able to promote health and longevity.</p

Analysis of transcription factor FOXO in the regulation of stress, aging, and neuromuscular tissue homeostasis in Drosophila melanogaster

The transcription factor FOXO is a known regulator of tissue homeostasis and animal lifespan. It was first identified in its ability to promote longevity through the insulin signaling pathway, and has since been implicated in numerous cellular processes. Members of the FOXO protein family control a wide array of cellular functions including metabolism, cell cycle arrest, apoptosis, stress resistance, and aging. In response to various signaling cues, FOXO proteins can localize into the nucleus and interact with DNA to regulate transcription. FOXO is known as a longevity gene, however, how FOXO behavior changes during aging is not well understood. Normal aging involves a progressive decline in cell function and an accumulation of oxidative damage. Additionally, with age comes a reduction in FOXO gene expression, and FOXO protein activity can become dysregulated. To resolve how FOXO activity changes with normal aging, we began with chromatin immunoprecipitation sequencing (ChIP-seq) to compare differences in FOXO chromatin binding between young and old organisms, using Drosophila melanogaster as a model. In Drosophila, there is only one homolog representing the FOXO protein family, known as dFOXO. Through our investigation, we found that the number of dFOXO-bound DNA regions decreases with age, and see a number of these targeted genes undergo changes in expression with aging. Some pathways targeted by FOXO at a young age are Hippo, WNT, and MAPK signaling pathways. FOXO is also known to mitigate oxidative stress, which is a contributing factor to age-related cellular degeneration. To understand FOXO dynamics in response to oxidative stress, we used Mass Spectrometry to evaluate dFOXO protein interacting networks under control and stress conditions. We observed a change in several dFOXO partners under paraquat-induced stress such as Stonewall, a chromatin modulator, and Hangover, a key transcription factor regulating neuromuscular junction (NMJ) morphology and neuronal activity. Both FOXO and Hangover show altered NMJ morphologies, and have potentially disrupted vesicle cycling. Given that FOXO has been linked to homeostatic maintenance of neuronal processes across animal species, we performed genetic analysis to investigate how dFOXO regulate NMJ aging in adult flies. Adult NMJs are known to undergo loss of synaptic homeostasis with aging, which causes functional decline and can lead to neurodegeneration. We profiled adult Drosophila abdominal NMJs and found that loss of function FOXO mutants exhibited morphological profiles similar to those of middle aged wild-type flies. We also observed an abnormal accumulation of late endosomes associated with the NMJ both in aged flies and with knockdown of motor neuron FOXO. Overexpression of FOXO can delay this accumulation, suggesting FOXO is a positive regulator of neuronal homeostasis. We performed a genetic screen and identified pathways such as MAPK that act downstream of FOXO to control NMJ homeostasis during aging. Collectively, this thesis provides evidence for global changes in FOXO chromatin binding activity under normal aging, highlights FOXO protein network dynamics that occur under oxidative stress, and displays homeostatic regulation of the adult Drosophila neuromuscular junction. These results illustrate how the transcription factor FOXO acts as a modulator of cellular homeostasis, and how FOXO activity is able to promote health and longevity