2022 Articles

Epigenetic aging of the demographically non-aging naked mole-rat


Kerepesi C, Meer MV, Ablaeva J, Amoroso VG, Lee SG, Zhang B, Gerashchenko MV, Trapp A, Yim SH, Lu AT, Levine ME, Seluanov A, Horvath S, Park TJ, Gorbunova V, Gladyshev VN.


Nature communications  

doi: 10.1038/s41467-022-27959-9


Abstract The naked mole-rat (NMR) is an exceptionally long-lived rodent that shows no increase of mortality with age, defining it as a demographically non-aging mammal. Here, we perform bisulfite sequencing of the blood of > 100 NMRs, assessing > 3 million common CpG sites. Unsupervised clustering based on sites whose methylation correlates with age reveals an age-related methylome remodeling, and we also observe a methylome information loss, suggesting that NMRs age. We develop an epigenetic aging clock that accurately predicts the NMR age. We show that these animals age much slower than mice and much faster than humans, consistent with their known maximum lifespans. Interestingly, patterns of age-related changes of clock sites in Tert and Prpf19 differ between NMRs and mice, but there are also sites conserved between the two species. Together, the data indicate that NMRs, like other mammals, epigenetically age even in the absence of demographic aging of this species. More Information


Rapamycin treatment during development extends life span and health span of male mice and Daphnia magna


Shindyapina AV, Cho Y, Kaya A, Tyshkovskiy A, Castro JP, Deik A, Gordevicius J, Poganik JR, Clish CB, Horvath S, Peshkin L, Gladyshev VN.


Science advances  

doi: 10.1126/sciadv.abo5482


Abstract Development is tightly connected to aging, but whether pharmacologically targeting development can extend life remains unknown. Here, we subjected genetically diverse UMHET3 mice to rapamycin for the first 45 days of life. The mice grew slower and remained smaller than controls for their entire lives. Their reproductive age was delayed without affecting offspring numbers. The treatment was sufficient to extend the median life span by 10%, with the strongest effect in males, and helped to preserve health as measured by frailty index scores, gait speed, and glucose and insulin tolerance tests. Mechanistically, the liver transcriptome and epigenome of treated mice were younger at the completion of treatment. Analogous to mice, rapamycin exposure during development robustly extended the life span of Daphnia magna and reduced its body size. Overall, the results demonstrate that short-term rapamycin treatment during development is a novel longevity intervention that acts by slowing down development and aging, suggesting that aging may be targeted already early in life. More Information


Emerging rejuvenation strategies-Reducing the biological age


Zhang B, Trapp A, Kerepesi C, Gladyshev VN.


Aging cell  

doi: 10.1111/acel.13538


Abstract Several interventions have recently emerged that were proposed to reverse rather than just attenuate aging, but the criteria for what it takes to achieve rejuvenation remain controversial. Distinguishing potential rejuvenation therapies from other longevity interventions, such as those that slow down aging, is challenging, and these anti-aging strategies are often referred to interchangeably. We suggest that the prerequisite for a rejuvenation intervention is a robust, sustained, and systemic reduction in biological age, which can be assessed by biomarkers of aging, such as epigenetic clocks. We discuss known and putative rejuvenation interventions and comparatively analyze them to explore underlying mechanisms. More Information


Selenocysteine Machinery Primarily Supports TXNRD1 and GPX4 Functions and Together They Are Functionally Linked with SCD and PRDX6


Santesmasses D, Gladyshev VN.


Biomolecules  

doi: 10.3390/biom12081049


Abstract The human genome has 25 genes coding for selenocysteine (Sec)-containing proteins, whose synthesis is supported by specialized Sec machinery proteins. Here, we carried out an analysis of the co-essentiality network to identify functional partners of selenoproteins and Sec machinery. One outstanding cluster included all seven known Sec machinery proteins and two critical selenoproteins, GPX4 and TXNRD1. Additionally, these nine genes were further positively associated with PRDX6 and negatively with SCD, linking the latter two genes to the essential role of selenium. We analyzed the essentiality scores of gene knockouts in this cluster across one thousand cancer cell lines and found that Sec metabolism genes are strongly selective for a subset of primary tissues, suggesting that certain cancer cell lineages are particularly dependent on selenium. A separate outstanding cluster included selenophosphate synthetase SEPHS1, which was linked to a group of transcription factors, whereas the remaining selenoproteins were linked neither to these clusters nor among themselves. The data suggest that key components of Sec machinery have already been identified and that their primary role is to support the functions of GPX4 and TXNRD1, with further functional links to PRDX6 and SCD. More Information


The selenoprotein methionine sulfoxide reductase B1 (MSRB1)


Tarrago L, Kaya A, Kim HY, Manta B, Lee BC, Gladyshev VN.


Free radical biology & medicine  

doi: 10.1016/j.freeradbiomed.2022.08.043


Abstract Methionine (Met) can be oxidized to methionine sulfoxide (MetO), which exist as R- and S-diastereomers. Present in all three domains of life, methionine sulfoxide reductases (MSR) are the enzymes that reduce MetO back to Met. Most characterized among them are MSRA and MSRB, which are strictly stereospecific for the S- and R-diastereomers of MetO, respectively. While the majority of MSRs use a catalytic Cys to reduce their substrates, some employ selenocysteine. This is the case of mammalian MSRB1, which was initially discovered as selenoprotein SELR or SELX and later was found to exhibit an MSRB activity. Genomic analyses demonstrated its occurrence in most animal lineages, and biochemical and structural analyses uncovered its catalytic mechanism. The use of transgenic mice and mammalian cell culture revealed its physiological importance in the protection against oxidative stress, maintenance of neuronal cells, cognition, cancer cell proliferation, and the immune response. Coincident with the discovery of Met oxidizing MICAL enzymes, recent findings of MSRB1 regulating the innate immunity response through reversible stereospecific Met-R-oxidation of cytoskeletal actin opened up new avenues for biological importance of MSRB1 and its role in disease. In this review, we discuss the current state of research on MSRB1, compare it with other animal Msrs, and offer a perspective on further understanding of biological functions of this selenoprotein. More Information


Selenoprotein TXNRD3 supports male fertility via the redox regulation of spermatogenesis


Dou Q, Turanov AA, Mariotti M, Hwang JY, Wang H, Lee SG, Paulo JA, Yim SH, Gygi SP, Chung JJ, Gladyshev VN.


The Journal of biological chemistry  

doi: 10.1016/j.jbc.2022.102183


Abstract Thioredoxin/glutathione reductase (TXNRD3) is a selenoprotein composed of thioredoxin reductase and glutaredoxin domains. This NADPH-dependent thiol oxidoreductase evolved through gene duplication within the Txnrd family, is expressed in the testes, and can reduce both thioredoxin and glutathione in vitro; however, the function of this enzyme remains unknown. To characterize the function of TXNRD3 in vivo, we generated a strain of mice bearing deletion of Txnrd3 gene. We show that these Txnrd3 knockout mice are viable and without discernable gross phenotypes, and also that TXNRD3 deficiency leads to fertility impairment in male mice. We found that Txnrd3 knockout animals exhibited a lower fertilization rate in vitro, a sperm movement phenotype, and an altered thiol redox status in sperm cells. Proteomic analyses further revealed a broad range of substrates reduced by TXNRD3 during sperm maturation, presumably as a part of sperm quality control. Taken together, these results show that TXNRD3 plays a critical role in male reproduction via the thiol redox control of spermatogenesis. More Information


In vivo cyclic induction of the FOXM1 transcription factor delays natural and progeroid aging phenotypes and extends healthspan


Ribeiro R, Macedo JC, Costa M, Ustiyan V, Shindyapina AV, Tyshkovskiy A, Gomes RN, Castro JP, Kalin TV, Vasques-Nóvoa F, Nascimento DS, Dmitriev SE, Gladyshev VN, Kalinichenko VV, Logarinho E.


Nature aging  

doi: 10.1038/s43587-022-00209-9


Abstract The FOXM1 transcription factor exhibits pleiotropic C-terminal transcriptional and N-terminal non-transcriptional functions in various biological processes critical for cellular homeostasis. We previously found that FOXM1 repression during cellular aging underlies the senescence phenotypes, which were vastly restored by overexpressing transcriptionally active FOXM1. Yet, it remains unknown whether increased expression of FOXM1 can delay organismal aging. Here, we show that in vivo cyclic induction of an N-terminal truncated FOXM1 transgene on progeroid and naturally aged mice offsets aging-associated repression of full-length endogenous Foxm1, reinstating both transcriptional and non-transcriptional functions. This translated into mitigation of several cellular aging hallmarks, as well as molecular and histopathological progeroid features of the short-lived Hutchison-Gilford progeria mouse model, significantly extending its lifespan. FOXM1 transgene induction also reinstated endogenous Foxm1 levels in naturally aged mice, delaying aging phenotypes while extending their lifespan. Thus, we disclose that FOXM1 genetic rewiring can delay senescence-associated progeroid and natural aging pathologies. More Information


Selenophosphate synthetase 1 deficiency exacerbates osteoarthritis by dysregulating redox homeostasis


Kang D, Lee J, Jung J, Carlson BA, Chang MJ, Chang CB, Kang SB, Lee BC, Gladyshev VN, Hatfield DL, Lee BJ, Kim JH.


Nature communications  

doi: 10.1038/s41467-022-28385-7


Abstract Aging and mechanical overload are prominent risk factors for osteoarthritis (OA), which lead to an imbalance in redox homeostasis. The resulting state of oxidative stress drives the pathological transition of chondrocytes during OA development. However, the specific molecular pathways involved in disrupting chondrocyte redox homeostasis remain unclear. Here, we show that selenophosphate synthetase 1 (SEPHS1) expression is downregulated in human and mouse OA cartilage. SEPHS1 downregulation impairs the cellular capacity to synthesize a class of selenoproteins with oxidoreductase functions in chondrocytes, thereby elevating the level of reactive oxygen species (ROS) and facilitating chondrocyte senescence. Cartilage-specific Sephs1 knockout in adult mice causes aging-associated OA, and augments post-traumatic OA, which is rescued by supplementation of N-acetylcysteine (NAC). Selenium-deficient feeding and Sephs1 knockout have synergistic effects in exacerbating OA pathogenesis in mice. Therefore, we propose that SEPHS1 is an essential regulator of selenium metabolism and redox homeostasis, and its dysregulation governs the progression of OA. More Information


A chromosome-level genome of Antechinus flavipes provides a reference for an Australian marsupial genus with male death after mating


Tian R, Han K, Geng Y, Yang C, Shi C, Thomas PB, Pearce C, Moffatt K, Ma S, Xu S, Yang G, Zhou X, Gladyshev VN, Liu X, Fisher DO, Chopin LK, Leiner NO, Baker AM, Fan G, Seim I.


Molecular ecology resources  

doi: 10.1111/1755-0998.13501


Abstract The 15 species of small carnivorous marsupials that comprise the genus Antechinus exhibit semelparity, a rare life-history strategy in mammals where synchronized death occurs after one breeding season. Antechinus males, but not females, age rapidly (demonstrate organismal senescence) during the breeding season and show promise as new animal models of ageing. Some antechinus species are also threatened or endangered. Here, we report a chromosome-level genome of a male yellow-footed antechinus Antechinus flavipes. The genome assembly has a total length of 3.2 Gb with a contig N50 of 51.8 Mb and a scaffold N50 of 636.7 Mb. We anchored and oriented 99.7% of the assembly on seven pseudochromosomes and found that repetitive DNA sequences occupy 51.8% of the genome. Draft genome assemblies of three related species in the subfamily Phascogalinae, two additional antechinus species (Antechinus argentus and A. arktos) and the iteroparous sister species Murexia melanurus, were also generated. Preliminary demographic analysis supports the hypothesis that climate change during the Pleistocene isolated species in Phascogalinae and shaped their population size. A transcriptomic profile across the A. flavipes breeding season allowed us to identify genes associated with aspects of the male die-off. The chromosome-level A. flavipes genome provides a steppingstone to understanding an enigmatic life-history strategy and a resource to assist the conservation of antechinuses. More Information


Biosensor-Linked Immunosorbent Assay for the Quantification of Methionine Oxidation in Target Proteins


Lee HM, Choi DW, Kim S, Lee A, Kim M, Roh YJ, Jo YH, Cho HY, Lee HJ, Lee SR, Tarrago L, Gladyshev VN, Kim JH, Lee BC.


ACS sensors  

doi: 10.1021/acssensors.1c01819


Abstract Methionine oxidation is involved in regulating the protein activity and often leads to protein malfunction. However, tools for quantitative analyses of protein-specific methionine oxidation are currently unavailable. In this work, we developed a biological sensor that quantifies oxidized methionine in the form of methionine-R-sulfoxide in target proteins. The biosensor “tpMetROG” consists of methionine sulfoxide reductase B (MsrB), circularly permuted yellow fluorescent protein (cpYFP), thioredoxin, and protein G. Protein G binds to the constant region of antibodies against target proteins, specifically capturing them. Then, MsrB reduces the oxidized methionine in these proteins, leading to cpYFP fluorescence changes. We assessed this biosensor for quantitative analysis of methionine-R-sulfoxide in various proteins, such as calmodulin, IDLO, LegP, Sacde, and actin. We further developed an immunosorbent assay using the biosensor to quantify methionine oxidation in specific proteins such as calmodulin in animal tissues. The biosensor-linked immunosorbent assay proves to be an indispensable tool for detecting methionine oxidation in a protein-specific manner. This is a versatile tool for studying the redox biology of methionine oxidation in proteins. More Information


SEPHS1: Its evolution, function and roles in development and diseases


Bang J, Kang D, Jung J, Yoo TJ, Shim MS, Gladyshev VN, Tsuji PA, Hatfield DL, Kim JH, Lee BJ.


Archives of biochemistry and biophysics  

doi: 10.1016/j.abb.2022.109426


Abstract Selenophosphate synthetase (SEPHS) was originally discovered in prokaryotes as an enzyme that catalyzes selenophosphate synthesis using inorganic selenium and ATP as substrates. However, in contrast to prokaryotes, two paralogs, SEPHS1 and SEPHS2, occur in many eukaryotes. Prokaryotic SEPHS, also known as SelD, contains either cysteine (Cys) or selenocysteine (Sec) in the catalytic domain. In eukaryotes, only SEPHS2 carries out selenophosphate synthesis and contains Sec at the active site. However, SEPHS1 contains amino acids other than Sec or Cys at the catalytic position. Phylogenetic analysis of SEPHSs reveals that the ancestral SEPHS contains both selenophosphate synthesis and another unknown activity, and that SEPHS1 lost the selenophosphate synthesis activity. The three-dimensional structure of SEPHS1 suggests that its homodimer is unable to form selenophosphate, but retains ATPase activity to produce ADP and inorganic phosphate. The most prominent function of SEPHS1 is that it is implicated in the regulation of cellular redox homeostasis. Deficiency of SEPHS1 leads to the disturbance in the expression of genes involved in redox homeostasis. Different types of reactive oxygen species (ROS) are accumulated in response to SEPHS deficiency depending on cell or tissue types. The accumulation of ROS causes pleiotropic effects such as growth retardation, apoptosis, DNA damage, and embryonic lethality. SEPHS1 deficiency in mouse embryos affects retinoic signaling and other related signaling pathways depending on the embryonal stage until the embryo dies at E11.5. Dysregulated SEPHS1 is associated with the pathogenesis of various diseases including cancer, Crohn’s disease, and osteoarthritis. More Information


Epigenetic clocks, aging, and cancer


Johnstone SE, Gladyshev VN, Aryee MJ, Bernstein BE.


Science (New York, N.Y.)  

doi: 10.1126/science.abn4009


Abstract Global methylation changes in aging cells affect cancer risk and tissue homeostasis. More Information


Characterization of naked mole-rat hematopoiesis reveals unique stem and progenitor cell patterns and neotenic traits


Emmrich S, Trapp A, Tolibzoda Zakusilo F, Straight ME, Ying AK, Tyshkovskiy A, Mariotti M, Gray S, Zhang Z, Drage MG, Takasugi M, Klusmann JH, Gladyshev VN, Seluanov A, Gorbunova V.


The EMBO journal  

doi: 10.15252/embj.2021109694


Abstract Naked mole rats (NMRs) are the longest-lived rodents yet their stem cell characteristics remain enigmatic. Here, we comprehensively mapped the NMR hematopoietic landscape and identified unique features likely contributing to longevity. Adult NMRs form red blood cells in spleen and marrow, which comprise a myeloid bias toward granulopoiesis together with decreased B-lymphopoiesis. Remarkably, youthful blood and marrow single-cell transcriptomes and cell compositions are largely maintained until at least middle age. Similar to primates, the primitive stem and progenitor cell (HSPC) compartment is marked by CD34 and THY1. Stem cell polarity is seen for Tubulin but not CDC42, and is not lost until 12 years of age. HSPC respiration rates are as low as in purified human stem cells, in concert with a strong expression signature for fatty acid metabolism. The pool of quiescent stem cells is higher than in mice, and the cell cycle of hematopoietic cells is prolonged. By characterizing the NMR hematopoietic landscape, we identified resilience phenotypes such as an increased quiescent HSPC compartment, absence of age-related decline, and neotenic traits likely geared toward longevity. More Information


Redox regulation by TXNRD3 during epididymal maturation underlies capacitation-associated mitochondrial activity and sperm motility in mice


Wang H, Dou Q, Jeong KJ, Choi J, Gladyshev VN, Chung JJ.


The Journal of biological chemistry  

doi: 10.1016/j.jbc.2022.102077


Abstract During epididymal transit, redox remodeling protects mammalian spermatozoa, preparing them for survival in the subsequent journey to fertilization. However, molecular mechanisms of redox regulation in sperm development and maturation remain largely elusive. In this study, we report that thioredoxin-glutathione reductase (TXNRD3), a thioredoxin reductase family member particularly abundant in elongating spermatids at the site of mitochondrial sheath formation, regulates redox homeostasis to support male fertility. Using Txnrd3-/- mice, our biochemical, ultrastructural, and live cell imaging analyses revealed impairments in sperm morphology and motility under conditions of TXNRD3 deficiency. We find that mitochondria develop more defined cristae during capacitation in wildtype sperm. Furthermore, we show that absence of TXNRD3 alters thiol redox status in both the head and tail during sperm maturation and capacitation, resulting in defective mitochondrial ultrastructure and activity under capacitating conditions. These findings provide insights into molecular mechanisms of redox homeostasis and bioenergetics during sperm maturation, capacitation, and fertilization. More Information