Search

Alterations in metabolism, sleep patterns, body composition and hormone status are all key features of aging. While the hypothalamus is a well-conserved brain region that controls these homeostatic and survival-related behaviors, little is known about the intrinsic features of hypothalamic aging. Here, we perform single-nuclei RNA sequencing of 40,064 hypothalamic nuclei from young and aged female mice. We identify cell type-specific signatures of aging in neuronal subtypes as well as astrocytes and microglia. We uncover changes in cell types critical for metabolic regulation and body composition and in an area of the hypothalamus linked to cognition. Our analysis also reveals an unexpected female-specific feature of hypothalamic aging: the master regulator of X inactivation, Xist, is elevated with age, particularly in hypothalamic neurons. Moreover, using machine learning, we show that levels of X chromosome genes and Xist itself, can accurately predict cellular age. This study identifies critical cell-specific changes of the aging hypothalamus in mammals and uncovers a potential marker of neuronal aging in females.

Retrotransposons are a class of transposable elements capable of self-replication and insertion into new genomic locations. Across species, the mobilization of retrotransposons in somatic cells has been suggested to contribute to the cell and tissue functional decline that occurs during aging. Retrotransposons are broadly expressed across cell types, and de novo insertions have been observed to correlate with tumorigenesis. However, the extent to which new retrotransposon insertions occur during normal aging and their effect on cellular and animal function remains understudied. Here, we use a single nucleus whole genome sequencing approach in Drosophila to directly test whether transposon insertions increase with age in somatic cells. Analyses of nuclei from thoraces and indirect flight muscles using a newly developed pipeline, Retrofind, revealed no significant increase in the number of transposon insertions with age. Despite this, reducing the expression of two different retrotransposons, 412 and Roo, extended lifespan, but did not alter indicators of health such as stress resistance. This suggests a key role for transposon expression and not insertion in regulating longevity. Transcriptomic analyses revealed similar changes to gene expression in 412 and Roo knockdown flies and highlighted changes to genes involved in proteolysis and immune function as potential contributors to the observed changes in longevity. Combined, our data show a clear link between retrotransposon expression and aging.

Retrotransposons are a class of transposable elements capable of self-replication and insertion into new genomic locations. Across species, the mobilization of retrotransposons in somatic cells has been suggested to contribute to the cell and tissue functional decline that occurs during aging. Retrotransposon expression generally increases with age, andde novoinsertions have been observed to occur during tumorigenesis. However, the extent to which new retrotransposon insertions occur during normal aging and their effect on cellular and animal function remains understudied. Here we use a single nucleus whole genome sequencing approach inDrosophilato directly test whether transposon insertions increase with age in somatic cells. Analyses of nuclei from thoraces and indirect flight muscles using a newly developed pipeline, Retrofind, revealed no significant increase in the number of transposon insertions with age. Despite this, reducing the expression of two different retrotransposons,412andRoo,extends lifespan, without increasing stress resistance. This suggests a key role for transposon expression and not insertion in regulating longevity. Transcriptomic analyses revealed similar changes to gene expression in412andRooknockdown flies and highlighted potential changes to genes involved in proteolysis and immune function as potential contributors to the observed changes in longevity. Combined, our data show a clear link between retrotransposon expression and aging.Author SummaryWith the onset of modern medicine, the average age of the population has significantly increased, leading to more individuals living with chronic health issues. Rather than treat each age-associated disorder individually, one approach to target multiple health concerns simultaneously might to be target aging itself. Genomic instability is a hallmark of aging cells that has been proposed to be a key contributor to age-associated cellular decline. Transposons are mobile genetic elements capable of inserting into new genomic locations, thus having the potential to increase genomic instability. Consistent with this, transposon expression generally increases with age. However, the extent to which transposon insertions accumulate to disrupt the genome of cells within aging individuals has remained an open question. We specifically answer this through single cell whole genome sequencing and find that transposon insertions do not increase with age. Even though insertions did not increase, the expression of transposons is linked to aging, as reducing the expression of individual transposons extended lifespan. Transcriptome studies of these long-lived flies revealed increased expression of genes linked to proteolysis genes and to functioning of the immune system. Our study therefore establishes transposon expression, and not insertion, as a critical contributor to animal aging.

Compositional and transcriptional changes in the hematopoietic system have been used as biomarkers of immunosenescence and aging. Here, we use single-cell RNA-sequencing to study the aging peripheral blood in mice and characterize the changes in cell-type composition and transcriptional profiles associated with age. We identified 17 clusters from a total of 14,588 single cells. We detected a general upregulation of antigen processing and presentation and chemokine signaling pathways and a downregulation of genes involved in ribosome pathways with age. In old peripheral blood, we also observed an increased percentage of cells expressing senescence markers (Cdkn1a, and Cdkn2a). In addition, we detected a cluster of activated T cells exclusively found in old blood, with lower expression of Cd28 and higher expression of Bcl2 and Cdkn2a, suggesting that the cells are senescent and resistant to apoptosis.

Cells respond to a myriad of stressors by senescing, acquiring stable growth arrest, morphologic and metabolic changes, and a senescence-associated-secretory-phenotype (SASP). The heterogeneity of senescent cells (SnCs) and their SASP is vast, yet poorly characterized. SnCs have diverse roles in health and disease and are therapeutically targetable, making characterization of SnCs and harmonization of their nomenclature a priority. The Cellular Senescence Network (SenNet), a NIH Common Fund initiative, will leverage emerging single cell and spatial-omics to identify and map SnCs in numerous organs across the lifespan of humans and mice. A common coordinate framework will integrate the data, using validated, standardized methods, creating public 4-dimensional SnC atlases. Key SenNet deliverables include development of innovative tools/technologies to detect SnCs, biomarker discovery, common annotations to describe SnCs and extensive public data sets. The goal is to comprehensively understand and map SnCs for diagnostic and therapeutic purposes to improve human health.

Cellular senescence is the irreversible cell cycle arrest in response to DNA damage. Because senescent cells accumulate with age and contribute to chronic inflammation, they are promising therapeutic targets for healthspan extension. The senescent phenotype can vary depending on cell type and on the specific insults that induce senescence. This variability is also reflected in the extensive remodeling of the genome organization within the nucleus of senescent cells. Here, we give an overview of the nuclear changes that occur in different forms of senescence, including changes to chromatin state and composition and to the three-dimensional organization of the genome, as well as alterations to the nuclear envelope and to the accessibility of repetitive genomic regions. Many of these changes are shared across all forms of senescence, implicating nuclear organization as a fundamental driver of the senescent state and of how senescent cells interact with the surrounding tissue.

The problem of three-dimensional (3D) chromosome structure inference from Hi-C data sets is important and challenging. While bulk Hi-C data sets contain contact information derived from millions of cells and can capture major structural features shared by the majority of cells in the sample, they do not provide information about local variability between cells. Single-cell Hi-C can overcome this problem, but contact matrices are generally very sparse, making structural inference more problematic. We have developed a Bayesian multiscale approach, named Structural Inference via Multiscale Bayesian Approach, to infer 3D structures of chromosomes from single-cell Hi-C while including the bulk Hi-C data and some regularization terms as a prior. We study the landscape of solutions for each single-cell Hi-C data set as a function of prior strength and demonstrate clustering of solutions using data from the same cell.

Cellular Senescence is a state of irreversible cell cycle arrest, and the accumulation of senescent cells contributes to age- related organismal decline. The detrimental effects of cellular senescence are due to the senescence associated secretory phenotype (SASP), an array of signaling molecules and growth factors secreted by senescent cells that contribute to the sterile inflammation associated with aging tissues. Recent studies, both in vivo and in vitro, have highlighted the heterogeneous nature of the senescence phenotype. In particular, single cell transcriptomics has revealed that Oncogene Induced Senescence (OIS) is characterized by the presence of subpopulations of cells expressing different SASP profiles. We have generated a comprehensive dataset via single-cell transcriptional profiling of genetically homogenous clonal cell lines from different forms of senescence, including OIS, Replicative Senescence (RS), and DNA Damage Induced Senescence (DDIS). We identified subpopulations of cells that are common to all three major forms of senescence and show that the expression profiles of these subpopulations are driven by markers formerly identified in individual forms of senescence. These common signatures are characterized by chromatin modifiers, inflammation, extracellular matrix remodeling, and Ribosomal protein expression. The expression patterns of these subpopulations recapitulate primary and secondary senescence, a phenomenon where a preexisting (primary) senescent cell induces senescence in a neighboring (secondary) cell through cell-to-cell contact. Since it is still unclear what type of senescence occurs in-vivo with age, it is important to know that the formation of primary and secondary populations is common to multiple types of senescence since this mechanism could help explain how senescent cells accumulate in aged organisms. Finally, we show that these subpopulations show differential susceptibility to the senolytic agent Navitoclax, suggesting that senolytic agents targeting the apoptotic pathways may be clearing only a subset of senescent cells based on their inflammatory profiles in-vivo.

Compositional and transcriptional changes in the hematopoietic system have been used as biomarkers of immunosenescence and aging. Here, we use single-cell RNA-sequencing to study the aging peripheral blood in mice, and characterize the changes in cell-type composition and transcriptional profiles associated with age. We identified 17 clusters from a total of 14,588 single cells. We detected a general upregulation of antigen processing and presentation and chemokine signaling pathways and a downregulation of genes involved in ribosome pathways with age. We also observed increased percentage of cells expressing markers of senescence, Cdkn1a and Cdkn2a, in old peripheral blood. In addition, we detected a cluster of activated T cells that are exclusively found in old blood, with lower expression of Cd28 and higher expression of Bcl2 and Cdkn2a, suggesting that the cells are senescent and resistant to apoptosis.

LINE-1 retrotransposons are known to be expressed in early development, in tumors and in the germline. Less is known about LINE-1 expression at the single cell level, especially outside the context of cancer. Because LINE-1 elements are present at a high copy number, many transcripts that are not driven by the LINE-1 promoter nevertheless terminate at the LINE-1 39 UTR. Thus, 39 targeted single cell RNA-seq datasets are not appropriate for studying LINE-1. However, 59 targeted single cell datasets provide an opportunity to analyze LINE-1 expression at the single cell level. Most LINE-1 copies are 59 truncated, and a transcript that contains the LINE-1 59 UTR as its 59 end is likely to have been transcribed from its promoter. We developed a method, L1-sc (LINE-1 expression for single cells), to quantify LINE-1 expression in 59 targeted 10x genomics single cell RNA-seq datasets. Our method confirms that LINE-1 expression is high in cancer cells, but low or absent from immune cells. We also find that LINE-1 expression is elevated in epithelial compared to immune cells outside of the context of cancer and that it is also elevated in neurons compared to glia in the mouse hippocampus.

Cellular senescence is characterized by an irreversible cell cycle arrest and a pro‐inflammatory senescence‐associated secretory phenotype (SASP), which is a major contributor to aging and age‐related diseases. Clearance of senescent cells has been shown to improve brain function in mouse models of neurodegenerative diseases. However, it is still unknown whether senescent cell clearance alleviates cognitive dysfunction during the aging process. To investigate this, we first conducted single‐nuclei and single‐cell RNA‐seq in the hippocampus from young and aged mice. We observed an age‐dependent increase in p16Ink4a senescent cells, which was more pronounced in microglia and oligodendrocyte progenitor cells and characterized by a SASP. We then aged INK‐ATTAC mice, in which p16Ink4a‐positive senescent cells can be genetically eliminated upon treatment with the drug AP20187 and treated them either with AP20187 or with the senolytic cocktail Dasatinib and Quercetin. We observed that both strategies resulted in a decrease in p16Ink4a exclusively in the microglial population, resulting in reduced microglial activation and reduced expression of SASP factors. Importantly, both approaches significantly improved cognitive function in aged mice. Our data provide proof‐of‐concept for senolytic interventions' being a potential therapeutic avenue for alleviating age‐associated cognitive impairment., Senescence is a major contributor to aging and age‐related diseases. However, it is still unknown whether senolytics impact on cognitive function during the aging process. We found that both pharmacogenetic clearance of p16Ink4a senescent cells or treatment with senolytic cocktail Dasatinib and Quercetin, reduced senescent microglia in the hippocampus and improved cognitive function in aged mice.

Cellular senescence is characterized by an irreversible cell cycle arrest as well as a pro-inflammatory phenotype, thought to contribute to aging and age-related diseases. Neutrophils have essential roles in inflammatory responses; however, in certain contexts their abundance is associated with a number of age-related diseases, including liver disease. The relationship between neutrophils and cellular senescence is not well understood. Here, we show that telomeres in non-immune cells are highly susceptible to oxidative damage caused by neighboring neutrophils. Neutrophils cause telomere dysfunction both in vitro and ex vivo in a ROS-dependent manner. In a mouse model of acute liver injury, depletion of neutrophils reduces telomere dysfunction and senescence. Finally, we show that senescent cells mediate the recruitment of neutrophils to the aged liver and propose that this may be a mechanism by which senescence spreads to surrounding cells. Our results suggest that interventions that counteract neutrophil-induced senescence may be beneficial during aging and age-related disease. publishedVersion

The study of the 3-dimensional (3D) structure of chromosomes – the largest macromolecules in biology – is one of the most challenging to date in structural biology. Here, we develop a novel representation of chromosomes, as sequences of shape letters from a finite shape alphabet, which provides a compact and efficient way to analyze ensembles of chromosome shape data, akin to the analysis of texts in a language by using letters. We construct a Chromosome Shape Alphabet (CSA) from an ensemble of chromosome 3D structures inferred from Hi-C data – via SIMBA3D or other methods – by segmenting curves based on topologically associating domains (TADs) boundaries, and by clustering all TADs’ 3D structures into groups of similar shapes. The median shapes of these groups, with some pruning and processing, form the Chromosome Shape Letters (CSLs) of the alphabet. We provide a proof-of-concept for these CSLs by reconstructing independent test curves using only CSLs (and corresponding transformations) and comparing these reconstructions with the original curves. Finally, we demonstrate how CSLs can be used to summarize the variability of shapes in an ensemble of chromosome 3D structures using generalized sequence logos.

Cellular senescence, an irreversible growth arrest triggered by a variety of stressors, plays important roles in normal physiology and tumor suppression, but accumulation of senescent cells with age contributes to the functional decline of tissues. Senescent cells undergo dramatic alterations to their chromatin landscape that affect genome accessibility and their transcriptional program. These include the loss of DNA-nuclear lamina interactions, the distension of centromeres, and changes in chromatin composition that can lead to the activation of retrotransposons. Here we discuss these findings, as well as recent advances in microscopy and genomics that have revealed the importance of the higher-order spatial organization of the genome in defining and maintaining the senescent state.

Oncogene-induced senescence (OIS) is a tumor suppressive response to oncogene activation that can be transmitted to neighboring cells through secreted factors of the senescence-associated secretory phenotype (SASP). Currently, primary and secondary senescent cells are not considered functionally distinct endpoints. Using single-cell analysis, we observed two distinct transcriptional endpoints, a primary endpoint marked by Ras and a secondary endpoint marked by Notch activation. We find that secondary oncogene-induced senescence in vitro and in vivo requires Notch, rather than SASP alone, as previously thought. Moreover, Notch signaling weakens, but does not abolish, SASP in secondary senescence. Global transcriptomic differences, a blunted SASP response, and the induction of fibrillar collagens in secondary senescence point toward a functional diversification between secondary and primary senescence.

Sea urchin embryos are excellent for in vivo functional studies because of their transparency and tractability in manipulation. They are also favorites for pharmacological approaches since they develop in an aquatic environment and addition of test substances is straightforward. A concern in many pharmacological tests though is the potential for pleiotropic effects that confound the conclusions drawn from the results. Precise cellular interpretations are often not feasible because the impact of the perturbant is not known. Here we use single-cell mRNA (messenger RNA) sequencing as a metric of cell types in the embryo and to determine the selectivity of two commonly used inhibitors, one each for the Wnt and the Delta-Notch pathways, on these nascent cell types. We identified 11 distinct cell types based on mRNA profiling, and that the cell lineages affected by Wnt and Delta/Notch inhibition were distinct from each other. These data support specificity and distinct effects of these signaling pathways in the embryo and illuminate how these conserved pathways selectively regulate cell lineages at a single cell level. Overall, we conclude that single cell RNA-seq analysis in this embryo is revealing of the cell types present during development, of the changes in the gene regulatory network resulting from inhibition of various signaling pathways, and of the selectivity of these pathways in influencing developmental trajectories.

Summary Mice deficient for SIRT6 exhibit a severely shortened lifespan, growth retardation, and highly elevated LINE1 (L1) activity. Here we report that SIRT6-deficient cells and tissues accumulate abundant cytoplasmic L1 cDNA, which triggers strong type I interferon response via activation of cGAS. Remarkably, nucleoside reverse-transcriptase inhibitors (NRTIs), which inhibit L1 retrotransposition, significantly improved health and lifespan of SIRT6 knockout mice and completely rescued type I interferon response. In tissue culture, inhibition of L1 with siRNA or NRTIs abrogated type I interferon response, in addition to a significant reduction of DNA damage markers. These results indicate that L1 activation contributes to the pathologies of SIRT6 knockout mice. Similarly, L1 transcription, cytoplasmic cDNA copy number, and type I interferons were elevated in the wild-type aged mice. As sterile inflammation is a hallmark of aging, we propose that modulating L1 activity may be an important strategy for attenuating age-related pathologies.

Oncogene induced senescence (OIS) is a tumour suppressive response to oncogene activation that can be transmitted to neighbouring cells through secreted factors of the senescence associated secretory phenotype (SASP). Using single-cell transcriptomics we observed two distinct endpoints, a primary marked by Ras and a secondary by Notch. We find that secondary senescence in vitro and in vivo requires Notch, rather than SASP alone as previously thought. Currently, primary and secondary senescent cells are not thought of as functionally distinct endpoints. A blunted SASP response and the induction of fibrillar collagens in secondary senescence compared to OIS point towards a functional diversification.One Sentence SummaryNotch signalling is an essential driver of secondary senescence with primary and secondary senescence being distinct molecular endpoints.

Retrotransposable elements are deleterious at many levels, and the failure of host surveillance systems for these elements can thus have negative consequences. However, the contribution of retrotransposon activity to ageing and age-associated diseases is not known. Here we show that during cellular senescence, L1 (also known as LINE-1) retrotransposable elements become transcriptionally derepressed and activate a type-I interferon (IFN-I) response. The IFN-I response is a phenotype of late senescence and contributes to the maintenance of the senescence-associated secretory phenotype. The IFN-I response is triggered by cytoplasmic L1 cDNA, and is antagonized by inhibitors of the L1 reverse transcriptase. Treatment of aged mice with the nucleoside reverse transcriptase inhibitor lamivudine downregulated IFN-I activation and age-associated inflammation (inflammaging) in several tissues. We propose that the activation of retrotransposons is an important component of sterile inflammation that is a hallmark of ageing, and that L1 reverse transcriptase is a relevant target for the treatment of age-associated disorders. During cellular senescence in human and mouse cells, L1 transposons become transcriptionally derepressed and trigger a type-1 interferon response, which contributes to age-associated inflammation and age-related phenotypes.

Transposons are genome components that account for the majority of genome size in many organisms, behaving as parasitic entities and interfering with the translation mechanism. Chromatin structure influences the activity of transposons, by coordinating genome accessibility for the expression and insertion of these sequences. As a case study, we show evidences of an evolutionary response of the chromatin structure to a variation in the activity of Long Interspersed Elements (LINEs) during mammals evolution, with focus on the murine radiation and primate evolution. LINEs activity was measured using a biophysical approach for modeling LINEs as an ecosystem, where different strains of transposons might reproduce, die and compete for access to the translational machinery of the host. The model, based on the discrete stochastic processes of amplification and deactivation of LINEs copies, has been adapted to the data using Bayesian statistics to estimate its main parameters: rate of growth of transposons copy number and rate of past competition between transposons variants. This approach allows to estimate the activity of ancient LINE strains still present in the genome as deactivated components, and the possible competition among different strains. We leverage these results to highlight how the change in the chromatin structure of the murine species seems to be following an increase of LINEs activity during the appearance of the murine specific strain Lx. On the contrary, a similar response is absent in primates evolution, which follows a decrease of LINEs activity during the amplication of primate specific LIMA/LPB strains.

SUMMARYMice deficient for SIRT6 exhibit a severely shortened lifespan, growth retardation, and highly elevated LINE1 (L1) activity. Here we report that SIRT6 deficient cells and tissues accumulate abundant cytoplasmic L1 cDNA which triggers massive type I interferon response via activation of cGAS. Remarkably, nucleoside reverse transcriptase inhibitors (NRTIs), which inhibit L1 retrotransposition, significantly improved health and lifespan of SIRT6 knockout mice and completely rescued type I interferon response. In tissue culture, inhibition of L1 with siRNA or NRTIs abrogated type I interferon response, in addition to a significant reduction of DNA damage markers. These results indicate that L1 activation contributes to the pathologies of SIRT6 knockout mice. Similarly, L1 transcription, cytoplasmic cDNA copy number and type I interferons were elevated in the wild type aged mice. As sterile inflammation is a hallmark of aging we propose that modulating L1 activity may be an important strategy for attenuating age-related pathologies.HighlightsSIRT6 KO mice accumulate L1 cDNA triggering type I interferon response via cGAS pathwayWild type aged mice accumulate L1 cDNA and display type I interferon responseReverse transcriptase inhibitors rescue type I interferon response and DNA damageReverse transcriptase inhibitors extend lifespan and improve health of SIRT6 KO mice

Recent advances in genomics and imaging technologies have increased our ability to interrogate the 3D conformation of chromosomes and to better understand principles of organization and dynamics, as well as how their alteration can lead to disease. In this review we describe how these technologies have shed new light into the role of the 3D organization of the genome in defining cellular states in aging and age-associated diseases. We compare the genomic organization in cellular senescence and cancer, discuss the role of the lamina in maintaining the structural and functional integrity of the genome, and we highlight the recent findings on how this organization breaks down in disease states.

Cell‐free DNA (cfDNA) is present in the circulating plasma and other body fluids and is known to originate mainly from apoptotic cells. Here, we provide the first in vivo evidence of global and local chromatin changes in human aging by analyzing cfDNA from the blood of individuals of different age groups. Our results show that nucleosome signals inferred from cfDNA are consistent with the redistribution of heterochromatin observed in cellular senescence and aging in other model systems. In addition, we detected a relative cfDNA loss at several genomic locations, such as transcription start and termination sites, 5′UTR of L1HS retrotransposons and dimeric AluY elements with age. Our results also revealed age and deteriorating health status correlate with increased enrichment of signals from cells in different tissues. In conclusion, our results show that the sequencing of circulating cfDNA from human blood plasma can be used as a noninvasive methodology to study age‐associated changes to the epigenome in vivo.

The problem of 3D chromosome structure inference from Hi-C datasets is important and challenging. While bulk Hi-C datasets contain contact information derived from millions of cells, and can capture major structural features shared by the majority of cells in the sample, they do not provide information about local variability between cells. Single cell Hi-C can overcome this problem, but contact matrices are generally very sparse, making structural inference more problematic. We have developed a Bayesian multiscale approach, named SIMBA3D, to infer 3D structures of chromosomes from single cell Hi-C while including the bulk Hi-C data and some regularization terms as a prior. We study the landscape of solutions for each single-cell Hi-C dataset as a function of prior strength and demonstrate clustering of solutions using data from the same cell.

Senescence is a cellular response triggered by diverse stresses. It can be beneficial, as a tumour suppressive response to oncogene activation, or detrimental as it drives pathology of aging. Senescence can be transmitted to neighbouring cells through secreted factors of the senescence associated secretory phenotype (SASP). Thus far, primary and secondary senescence have been considered identical phenotypes. We used single-cell transcriptomics in co-culture systems to decipher heterogeneity between primary and secondary Ras-induced senescence and observed two distinct transcriptional trajectories, one marked by Ras and the other by Notch. Furthermore, we find that secondary senescence in vitro and in vivo is driven by Notch, rather than by the SASP as previously thought. Our approach demonstrates the power of single-cell approaches in interrogation of complex biological systems. Based on these results, we conclude that primary senescence and secondary senescence are distinct molecular endpoints, and secondary senescence is driven by juxtacrine notch signalling, not SASP.

Previous research has shown that gold nanoparticles can increase the effectiveness of radiation on cancer cells. Improved radiation effectiveness would allow lower radiation doses given to patients, reducing adverse effects; alternatively, it would provide more cancer killing at current radiation doses. Damage from radiation and gold nanoparticles depends in part on the Auger effect, which is very localized; thus, it is important to place the gold nanoparticles on or in the cancer cells. In this work, we use the pH-sensitive, tumor-targeting agent, pH Low-Insertion Peptide (pHLIP), to tether 1.4-nm gold nanoparticles to cancer cells. We find that the conjugation of pHLIP to gold nanoparticles increases gold uptake in cells compared with gold nanoparticles without pHLIP, with the nanoparticles distributed mostly on the cellular membranes. We further find that gold nanoparticles conjugated to pHLIP produce a statistically significant decrease in cell survival with radiation compared with cells without gold nanoparticles and cells with gold alone. In the context of our previous findings demonstrating efficient pHLIP-mediated delivery of gold nanoparticles to tumors, the obtained results serve as a foundation for further preclinical evaluation of dose enhancement.

SummaryMYC is a highly pleiotropic transcription factor whose deregulation promotes cancer. In contrast, we find that Myc haploinsufficient (Myc+/–) mice exhibit increased lifespan. They show resistance to several age-associated pathologies, including osteoporosis, cardiac fibrosis, and immunosenescence. They also appear to be more active, with a higher metabolic rate and healthier lipid metabolism. Transcriptomic analysis reveals a gene expression signature enriched for metabolic and immune processes. The ancestral role of MYC as a regulator of ribosome biogenesis is reflected in reduced protein translation, which is inversely correlated with longevity. We also observe changes in nutrient and energy sensing pathways, including reduced serum IGF-1, increased AMPK activity, and decreased AKT, TOR, and S6K activities. In contrast to observations in other longevity models, Myc+/– mice do not show improvements in stress management pathways. Our findings indicate that MYC activity has a significant impact on longevity and multiple aspects of mammalian healthspan.PaperClip