Comparative analysis of animal lifespan.

Comparative studies of aging are a promising approach to identifying general properties of and processes leading to aging. While to date, many comparative studies of aging in animals have focused on relatively narrow species groups, methodological innovations now allow for studies that include evolutionary distant species. However, comparative studies of aging across a wide range of species that have distinct life histories introduce additional challenges in experimental design. Here, we discuss these challenges, highlight the most pressing problems that need to be solved, and provide suggestions based on current approaches to successfully carry out comparative aging studies across the animal kingdom.

Modeling collective cell behavior in cancer: Perspectives from an interdisciplinary conversation.

Collective cell behavior contributes to all stages of cancer progression. Understanding how collective behavior emerges through cell-cell interactions and decision-making will advance our understanding of cancer biology and provide new therapeutic approaches. Here, we summarize an interdisciplinary discussion on multicellular behavior in cancer, draw lessons from other scientific disciplines, and identify future directions.

Functions of HP1 proteins in transcriptional regulation.

In eukaryotes, DNA is packaged into chromatin, which presents significant barriers to transcription. Non-histone chromatin proteins such as the Heterochromatin Protein 1 (HP1) proteins are critical regulators of transcription, contributing to gene regulation through a variety of molecular mechanisms. HP1 proteins are highly conserved, and many eukaryotic genomes contain multiple HP1 genes. Given the presence of multiple HP1 family members within a genome, HP1 proteins can have unique as well as shared functions. Here, we review the mechanisms by which HP1 proteins contribute to the regulation of transcription. Focusing on the Drosophila melanogaster HP1 proteins, we examine the role of these proteins in regulating the transcription of genes, transposable elements, and piRNA clusters. In D. melanogaster, as in other species, HP1 proteins can act as transcriptional repressors and activators. The available data reveal that the precise impact of HP1 proteins on gene expression is highly context dependent, on the specific HP1 protein involved, on its protein partners present, and on the specific chromatin context the interaction occurs in. As a group, HP1 proteins utilize a variety of mechanisms to contribute to transcriptional regulation, including both transcriptional (i.e. chromatin-based) and post-transcriptional (i.e. RNA-based) processes. Despite extensive studies of this important protein family, open questions regarding their functions in gene regulation remain, specifically regarding the role of hetero- versus homodimerization and post-translational modifications of HP1 proteins.

Sex-specific aging in animals: Perspective and future directions.

Sex differences in aging occur in many animal species, and they include sex differences in lifespan, in the onset and progression of age-associated decline, and in physiological and molecular markers of aging. Sex differences in aging vary greatly across the animal kingdom. For example, there are species with longer-lived females, species where males live longer, and species lacking sex differences in lifespan. The underlying causes of sex differences in aging remain mostly unknown. Currently, we do not understand the molecular drivers of sex differences in aging, or whether they are related to the accepted hallmarks or pillars of aging or linked to other well-characterized processes. In particular, understanding the role of sex-determination mechanisms and sex differences in aging is relatively understudied. Here, we take a comparative, interdisciplinary approach to explore various hypotheses about how sex differences in aging arise. We discuss genomic, morphological, and environmental differences between the sexes and how these relate to sex differences in aging. Finally, we present some suggestions for future research in this area and provide recommendations for promising experimental designs.

The Epigenome and Beyond: How Does Non-genetic Inheritance Change Our View of Evolution?

Evidence from across the tree of life suggests that epigenetic inheritance is more common than previously thought. If epigenetic inheritance is indeed as common as the data suggest, this finding has potentially important implications for evolutionary theory and our understanding of how evolution and adaptation progress. However, we currently lack an understanding of how common various epigenetic inheritance types are, and how they impact phenotypes. In this perspective, we review the open questions that need to be addressed to fully integrate epigenetic inheritance into evolutionary theory and to develop reliable predictive models for phenotypic evolution. We posit that addressing these challenges will require the collaboration of biologists from different disciplines and a focus on the exploration of data and phenomena without preconceived limits on potential mechanisms or outcomes.

The Drosophila HP1 family is associated with active gene expression across chromatin contexts.

Drosophila Heterochromatin Protein 1a (HP1a) is essential for heterochromatin formation and is involved in transcriptional silencing. However, certain loci require HP1a to be transcribed. One model posits that HP1a acts as a transcriptional silencer within euchromatin while acting as an activator within heterochromatin. However, HP1a has been observed as an activator of a set of euchromatic genes. Therefore, it is not clear whether, or how, chromatin context informs the function of HP1 proteins. To understand the role of HP1 proteins in transcription, we examined the genome-wide binding profile of HP1a as well as two other Drosophila HP1 family members, HP1B and HP1C, to determine whether coordinated binding of these proteins is associated with specific transcriptional outcomes. We found that HP1 proteins share many of their endogenous binding targets. These genes are marked by active histone modifications and are expressed at higher levels than nontarget genes in both heterochromatin and euchromatin. In addition, HP1 binding targets displayed increased RNA polymerase pausing compared with nontarget genes. Specifically, colocalization of HP1B and HP1C was associated with the highest levels of polymerase pausing and gene expression. Analysis of HP1 null mutants suggests these proteins coordinate activity at transcription start sites to regulate transcription. Depletion of HP1B or HP1C alters expression of protein-coding genes bound by HP1 family members. Our data broaden understanding of the mechanism of transcriptional activation by HP1a and highlight the need to consider particular protein-protein interactions, rather than broader chromatin context, to predict impacts of HP1 at transcription start sites.

Exercise-induced changes in climbing performance.

Exercise is recommended to promote health and prevent a range of diseases. However, how exercise precipitates these benefits is unclear, nor do we understand why exercise responses differ so widely between individuals. We investigate how climbing ability in Drosophila melanogaster changes in response to an exercise treatment. We find extensive variation in baseline climbing ability and exercise-induced changes ranging from -13% to +20% in climbing ability. Climbing ability, and its exercise-induced change, is sex- and genotype-dependent. GWASs implicate 'cell-cell signalling' genes in the control of climbing ability. We also find that animal activity does not predict climbing ability and that the exercise-induced climbing ability change cannot be predicted from the activity level induced by the exercise treatment. These results provide promising new avenues for further research into the molecular pathways controlling climbing activity and illustrate the complexities involved in trying to predict individual responses to exercise.

GWAS reveal a role for the central nervous system in regulating weight and weight change in response to exercise.

Body size and weight show considerable variation both within and between species. This variation is controlled in part by genetics, but also strongly influenced by environmental factors including diet and the level of activity experienced by the individual. Due to the increasing obesity epidemic in much of the world, there is considerable interest in the genetic factors that control body weight and how weight changes in response to exercise treatments. Here, we address this question in the Drosophila model system, utilizing 38 strains of the Drosophila Genetics Reference Panel. We use GWAS to identify the molecular pathways that control weight and weight changes in response to exercise. We find that there is a complex set of molecular pathways controlling weight, with many genes linked to the central nervous system (CNS). The CNS also plays a role in the weight change with exercise, in particular, signaling from the CNS. Additional analyses revealed that weight in Drosophila is driven by two factors, animal size, and body composition, as the amount of fat mass versus lean mass impacts the density. Thus, while the CNS appears to be important for weight and exercise-induced weight change, signaling pathways are particularly important for determining how exercise impacts weight.

Variation in the response to exercise stimulation in Drosophila: marathon runner versus sprinter genotypes.

Animals' behaviors vary in response to their environment, both biotic and abiotic. These behavioral responses have significant impacts on animal survival and fitness, and thus, many behavioral responses are at least partially under genetic control. In Drosophila, for example, genes impacting aggression, courtship behavior, circadian rhythms and sleep have been identified. Animal activity also is influenced strongly by genetics. My lab previously has used the Drosophila melanogaster Genetics Reference Panel (DGRP) to investigate activity levels and identified over 100 genes linked to activity. Here, I re-examined these data to determine whether Drosophila strains differ in their response to rotational exercise stimulation, not simply in the amount of activity, but in activity patterns and timing of activity. Specifically, I asked whether there are fly strains exhibiting either a 'marathoner' pattern of activity, i.e. remaining active throughout the 2 h exercise period, or a 'sprinter' pattern, i.e. carrying out most of the activity early in the exercise period. The DGRP strains examined differ significantly in how much activity is carried out at the beginning of the exercise period, and this pattern is influenced by both sex and genotype. Interestingly, there was no clear link between the activity response pattern and lifespan of the animals. Using genome-wide association studies (GWAS), I identified 10 high confidence candidate genes that control the degree to which Drosophila exercise behaviors fit a marathoner or sprinter activity pattern. This finding suggests that, similar to other aspects of locomotor behavior, the timing of activity patterns in response to exercise stimulation is under genetic control.

Epigenetics and genome stability.

Maintaining genome stability is essential to an organism's health and survival. Breakdown of the mechanisms protecting the genome and the resulting genome instability are an important aspect of the aging process and have been linked to diseases such as cancer. Thus, a large network of interconnected pathways is responsible for ensuring genome integrity in the face of the continuous challenges that induce DNA damage. While these pathways are diverse, epigenetic mechanisms play a central role in many of them. DNA modifications, histone variants and modifications, chromatin structure, and non-coding RNAs all carry out a variety of functions to ensure that genome stability is maintained. Epigenetic mechanisms ensure the functions of centromeres and telomeres that are essential for genome stability. Epigenetic mechanisms also protect the genome from the invasion by transposable elements and contribute to various DNA repair pathways. In this review, we highlight the integral role of epigenetic mechanisms in the maintenance of genome stability and draw attention to issues in need of further study.

Genetic Networks Underlying Natural Variation in Basal and Induced Activity Levels in Drosophila melanogaster.

Exercise is recommended by health professionals across the globe as part of a healthy lifestyle to prevent and/or treat the consequences of obesity. While overall, the health benefits of exercise and an active lifestyle are well understood, very little is known about how genetics impacts an individual's inclination for and response to exercise. To address this knowledge gap, we investigated the genetic architecture underlying natural variation in activity levels in the model system Drosophila melanogaster Activity levels were assayed in the Drosophila Genetics Reference Panel fly strains at baseline and in response to a gentle exercise treatment using the Rotational Exercise Quantification System. We found significant, sex-dependent variation in both activity measures and identified over 100 genes that contribute to basal and induced exercise activity levels. This gene set was enriched for genes with functions in the central nervous system and in neuromuscular junctions and included several candidate genes with known activity phenotypes such as flightlessness or uncoordinated movement. Interestingly, there were also several chromatin proteins among the candidate genes, two of which were validated and shown to impact activity levels. Thus, the study described here reveals the complex genetic architecture controlling basal and exercise-induced activity levels in D. melanogaster and provides a resource for exercise biologists.

New opportunities: Drosophila as a model system for exercise research.

Because of the growing rates of obesity in much of the world, exercise as a treatment option for obesity and as part of a healthy lifestyle is of great interest to the general public, health policy makers, and scientists alike. Despite the long history of exercise promotion and exercise research, there are still significant gaps in our understanding of how exercise impacts individuals and what role genetics plays in determining an individual's response to exercise. Model organisms are positioned uniquely to help address these questions because of the challenges associated with carrying out large-scale, well-controlled studies in humans. The fruit fly model system, Drosophila melanogaster, has joined the models used for exercise research only recently but already has made significant contributions to the field. In this review, we highlight the opportunities for exercise research in Drosophila. We review the resources available to researchers interested in using Drosophila for exercise research, focusing on the existing systems to induce exercise in Drosophila, to measure the amount of exercise performed, and to assess physical fitness. We illustrate the potential of the Drosophila system by drawing attention to pioneering studies in Drosophila exercise research and emphasize the unique opportunities this model system represents.

The Drosophila Dot Chromosome: Where Genes Flourish Amidst Repeats.

The F element of the Drosophila karyotype (the fourth chromosome in Drosophila melanogaster) is often referred to as the "dot chromosome" because of its appearance in a metaphase chromosome spread. This chromosome is distinct from other Drosophila autosomes in possessing both a high level of repetitious sequences (in particular, remnants of transposable elements) and a gene density similar to that found in the other chromosome arms, ∼80 genes distributed throughout its 1.3-Mb "long arm." The dot chromosome is notorious for its lack of recombination and is often neglected as a consequence. This and other features suggest that the F element is packaged as heterochromatin throughout. F element genes have distinct characteristics (e.g, low codon bias, and larger size due both to larger introns and an increased number of exons), but exhibit expression levels comparable to genes found in euchromatin. Mapping experiments show the presence of appropriate chromatin modifications for the formation of DNaseI hypersensitive sites and transcript initiation at the 5' ends of active genes, but, in most cases, high levels of heterochromatin proteins are observed over the body of these genes. These various features raise many interesting questions about the relationships of chromatin structures with gene and chromosome function. The apparent evolution of the F element as an autosome from an ancestral sex chromosome also raises intriguing questions. The findings argue that the F element is a unique chromosome that occupies its own space in the nucleus. Further study of the F element should provide new insights into chromosome structure and function.

HP1B is a euchromatic Drosophila HP1 homolog with links to metabolism.

Heterochromatin Protein 1 (HP1) proteins are an important family of chromosomal proteins conserved among all major eukaryotic lineages. While HP1 proteins are best known for their role in heterochromatin, many HP1 proteins function in euchromatin as well. As a group, HP1 proteins carry out diverse functions, playing roles in the regulation of gene expression, genome stability, chromatin structure, and DNA repair. While the heterochromatic HP1 proteins are well studied, our knowledge of HP1 proteins with euchromatic distribution is lagging behind. We have created the first mutations in HP1B, a Drosophila HP1 protein with euchromatic function, and the Drosophila homolog most closely related to mammalian HP1α, HP1ß, and HP1γ. We find that HP1B is a non-essential protein in Drosophila, with mutations affecting fertility and animal activity levels. In addition, animals lacking HP1B show altered food intake and higher body fat levels. Gene expression analysis of animals lacking HP1B demonstrates that genes with functions in various metabolic processes are affected primarily by HP1B loss. Our findings suggest that there is a link between the chromatin protein HP1B and the regulation of metabolism.

Measuring Exercise Levels in Drosophila melanogaster Using the Rotating Exercise Quantification System (REQS).

Drosophila melanogaster is a new model organism for studies in exercise biology. To date, two main exercise systems, the Power Tower and the Treadwheel have been described. However, a method to measure the amount of additional animal activity induced through the exercise treatment has been lacking. The Rotating Exercise Quantification System (REQS) fills this need, providing a measure of animal activity for animals experiencing rotational exercise. This protocol details how to use the REQS to assess animal activity during rotational exercise and illustrates the type of data that can be generated. Here, we demonstrate how the REQS is used to measure sex- and strain-specific differences in exercise induced activity. The REQS can also be used to evaluate the impact of various other experimental parameters such as age, diet, or population size on exercise induced activity. In addition, it can be used to compare the efficacy of different exercise training protocols. Importantly, it provides an opportunity to standardize exercise treatments between strains, allowing the researcher to achieve equal amounts of activity between groups if needed. Thus, the REQS is a notable new resource for exercise biologists working with the Drosophila model system and complements existing exercise systems.

Sex Differences in Aging: Genomic Instability.

Aging is characterized by decreasing physiological integration, reduced function, loss of resilience, and increased risk of death. Paradoxically, although women live longer, they suffer greater morbidity particularly late in life. These sex differences in human lifespan and healthspan are consistently observed in all countries and during every era for which reliable data exist. While these differences are ubiquitous in humans, evidence of sex differences in longevity and health for other species is more equivocal. Among fruit flies, nematodes, and mice, sex differences in lifespan vary depending on strain and treatment. In this review, we focus on sex differences in age-related alterations in DNA damage and mutation rates, telomere attrition, epigenetics, and nuclear architecture. We find that robust sex differences exist, eg, the higher incidence of DNA damage in men compared to women, but sex differences are not often conserved between species. For most mechanisms reviewed here, there are insufficient data to make a clear determination regarding the impact of sex, largely because sex differences have not been analyzed. Overall, our findings reveal an urgent need for well-designed studies that explicitly examine sex differences in molecular drivers of aging.

Characterization of the Rotating Exercise Quantification System (REQS), a novel Drosophila exercise quantification apparatus.

Obesity is a disease that has reached epidemic proportions in the United States and has prompted international legislation in an attempt to curtail its prevalence. Despite the fact that one of the most prescribed treatment options for obesity is exercise, the genetic mechanisms underlying exercise response in individuals are still largely unknown. The fruit fly Drosophila melanogaster is a promising new model for studying exercise genetics. Currently, the lack of an accurate method to quantify the amount of exercise performed by the animals is limiting the utility of the Drosophila model for exercise genetics research. To address this limitation, we developed the Rotational Exercise Quantification System (REQS), a novel apparatus that is able to simultaneously induce exercise in flies while recording their activity levels. Thus, the REQS provides a method to standardize Drosophila exercise and ensure that all animals irrespective of genotype and sex experience the same level of exercise. Here, we provide a basic characterization of the REQS, validate its measurements using video-tracking technology, illustrate its potential use by presenting a comparison of two different exercise regimes, and demonstrate that it can be used to detect genotype-dependent variation in activity levels.

The TreadWheel: A Novel Apparatus to Measure Genetic Variation in Response to Gently Induced Exercise for Drosophila.

Obesity is one of the dramatic health issues affecting developed and developing nations, and exercise is a well-established intervention strategy. While exercise-by-genotype interactions have been shown in humans, overall little is known. Using the natural negative geotaxis of Drosophila melanogaster, an important model organism for the study of genetic interactions, a novel exercise machine, the TreadWheel, can be used to shed light on this interaction. The mechanism for inducing exercise with the TreadWheel is inherently gentle, thus minimizing possible confounding effects of other stressors. Using this machine, we were able to assess large cohorts of adult flies from eight genetic lines for their response to exercise after one week of training. We measured their triglyceride, glycerol, protein, glycogen, glucose content, and body weight, as well as their climbing ability and feeding behavior in response to exercise. Exercised flies showed decreased stored triglycerides, glycogen, and body weight, and increased stored protein and climbing ability. In addition to demonstrating an overall effect of TreadWheel exercise on flies, we found significant interactions of exercise with genotype, sex, or genotype-by-sex effects for most of the measured phenotypes. We also observed interaction effects between exercise, genotype, and tissue (abdomen or thorax) for metabolite profiles, and those differences can be partially linked to innate differences in the flies' persistence in maintaining activity during exercise bouts. In addition, we assessed gene expression levels for a panel of 13 genes known to be associated with respiratory fitness and found that many responded to exercise. With this study, we have established the TreadWheel as a useful tool to study the effect of exercise in flies, shown significant genotype-specific and sex-specific impacts of exercise, and have laid the ground work for more extensive studies of how genetics, sex, environment, and aging interact with exercise to influence metabolic fitness in Drosophila.

Rho Kinase Inhibition as a Therapeutic for Progressive Supranuclear Palsy and Corticobasal Degeneration.

Progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD) are neurodegenerative four-repeat tauopathies with no cure. Mitigating pathogenic tau levels is a rational strategy for tauopathy treatment, but therapeutic targets with clinically available drugs are lacking. Here, we report that protein levels of the Rho-associated protein kinases (ROCK1 and ROCK2), p70 S6 kinase (S6K), and mammalian target of rapamycin (mTOR) were increased in PSP and CBD brains. RNAi depletion of ROCK1 or ROCK2 reduced tau mRNA and protein level in human neuroblastoma cells. However, additional phenotypes were observed under ROCK2 knockdown, including decreased S6K and phosphorylated mTOR levels. Pharmacologic inhibition of Rho kinases in neurons diminished detergent-soluble and -insoluble tau through a combination of autophagy enhancement and tau mRNA reduction. Fasudil, a clinically approved ROCK inhibitor, suppressed rough eye phenotype and mitigated pathogenic tau levels by inducing autophagic pathways in a Drosophila model of tauopathy. Collectively, these findings highlight the Rho kinases as rational therapeutic targets to combat tau accumulation in PSP and CBD. SIGNIFICANCE STATEMENT: Studies of progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD) suggest that mitigating pathogenic tau levels is a rational strategy for tauopathy treatment. In this report, the Rho-associated protein kinases (ROCK1 and ROCK2) are identified as novel drug targets for PSP and CBD. We show that elevated insoluble tau levels are associated with increased ROCK1 and ROCK2 in PSP and CBD brains, whereas experiments in cellular and animal models identify pharmacologic inhibition of ROCKs as a mechanism-based approach to reduce tau levels. Our study correlates bona fide changes in PSP and CBD brains with cellular models, identifies drug targets, and tests the therapeutic in vivo.