The analysis of pendolino (peo) mutants reveals differences in the fusigenic potential among Drosophila telomeres

Drosophila telomeres are sequence-independent structures that are maintained by transposition to chromosome ends of three specialized retroelements (HeT-A, TART and TAHRE; collectively designated as HTT) rather than telomerase activity. Fly telomeres are protected by the terminin complex (HOAP-HipHop-Moi-Ver) that localizes and functions exclusively at telomeres and by non-terminin proteins that do not serve telomere-specific functions. Although all Drosophila telomeres terminate with HTT arrays and are capped by terminin, they differ in the type of subtelomeric chromatin; the Y, XR, and 4L HTT are juxtaposed to constitutive heterochromatin, while the XL, 2L, 2R, 3L and 3R HTT are linked to the TAS repetitive sequences; the 4R HTT is associated with a chromatin that has features common to both euchromatin and heterochromatin. Here we show that mutations in pendolino (peo) cause telomeric fusions (TFs). The analysis of several peo mutant combinations showed that these TFs preferentially involve the Y, XR and 4th chromosome telomeres, a TF pattern never observed in the other 10 telomere-capping mutants so far characterized. peo encodes a non-terminin protein homologous to the E2 variant ubiquitin-conjugating enzymes. The Peo protein directly interacts with the terminin components, but peo mutations do not affect telomeric localization of HOAP, Moi, Ver and HP1a, suggesting that the peodependent telomere fusion phenotype is not due to loss of terminin from chromosome ends. peo mutants are also defective in DNA replication and PCNA recruitment. However, our results suggest that general defects in DNA replication are unable to induce TFs in Drosophila cells. We thus hypothesize that DNA replication in Peodepleted cells results in specific fusigenic lesions concentrated in heterochromatinassociated telomeres. Alternatively it is possible that Peo plays a dual function being independently required for DNA replication and telomere capping

Spatially resolved transcriptomics reveals innervation-responsive functional clusters in skeletal muscle

Striated muscle is a highly organized structure composed of well-defined anatomical domains with integrated but distinct assignments. So far, the lack of a direct correlation between tissue architecture and gene expression has limited our understanding of how each unit responds to physio-pathologic contexts. Here, we show how the combined use of spatially resolved transcriptomics and immunofluorescence can bridge this gap by enabling the unbiased identification of such domains and the characterization of their response to external perturbations. Using a spatiotemporal analysis, we follow changes in the transcriptome of specific domains in muscle in a model of denervation. Furthermore, our approach enables us to identify the spatial distribution and nerve dependence of atrophic signaling pathway and polyamine metabolism to glycolytic fibers. Indeed, we demonstrate that perturbations of polyamine pathway can affect muscle function. Our dataset serves as a resource for future studies of the mechanisms underlying skeletal muscle homeostasis and innervation

Ablations of Ghrelin and Ghrelin Receptor Exhibit Differential Metabolic Phenotypes and Thermogenic Capacity during Aging

mice are adaptive. mice.Our data therefore suggest that GHS-R ablation activates adaptive thermogenic function(s) in BAT and increases EE, thereby enabling the retention of a lean phenotype. This is the first direct evidence that the ghrelin signaling pathway regulates fat-burning BAT to affect energy balance during aging. This regulation is likely mediated through an as-yet-unidentified new ligand of GHS-R

Widespread horse-based mobility arose around 2200 BCE in Eurasia.

Horses revolutionized human history with fast mobility1. However, the timeline between their domestication and their widespread integration as a means of transport remains contentious2-4. Here we assemble a collection of 475 ancient horse genomes to assess the period when these animals were first reshaped by human agency in Eurasia. We find that reproductive control of the modern domestic lineage emerged around 2200 BCE, through close-kin mating and shortened generation times. Reproductive control emerged following a severe domestication bottleneck starting no earlier than approximately 2700 BCE, and coincided with a sudden expansion across Eurasia that ultimately resulted in the replacement of nearly every local horse lineage. This expansion marked the rise of widespread horse-based mobility in human history, which refutes the commonly held narrative of large horse herds accompanying the massive migration of steppe peoples across Europe around 3000 BCE and earlier3,5. Finally, we detect significantly shortened generation times at Botai around 3500 BCE, a settlement from central Asia associated with corrals and a subsistence economy centred on horses6,7. This supports local horse husbandry before the rise of modern domestic bloodlines

Fatality rate and predictors of mortality in an Italian cohort of hospitalized COVID-19 patients

Clinical features and natural history of coronavirus disease 2019 (COVID-19) differ widely among different countries and during different phases of the pandemia. Here, we aimed to evaluate the case fatality rate (CFR) and to identify predictors of mortality in a cohort of COVID-19 patients admitted to three hospitals of Northern Italy between March 1 and April 28, 2020. All these patients had a confirmed diagnosis of SARS-CoV-2 infection by molecular methods. During the study period 504/1697 patients died; thus, overall CFR was 29.7%. We looked for predictors of mortality in a subgroup of 486 patients (239 males, 59%; median age 71 years) for whom sufficient clinical data were available at data cut-off. Among the demographic and clinical variables considered, age, a diagnosis of cancer, obesity and current smoking independently predicted mortality. When laboratory data were added to the model in a further subgroup of patients, age, the diagnosis of cancer, and the baseline PaO2/FiO2 ratio were identified as independent predictors of mortality. In conclusion, the CFR of hospitalized patients in Northern Italy during the ascending phase of the COVID-19 pandemic approached 30%. The identification of mortality predictors might contribute to better stratification of individual patient risk

Telomere fusion in Drosophila: The role of subtelomeric chromatin

Drosophila telomeres are maintained by transposition to chromosome ends of the HeT-A, TART and TAHRE retrotransposons, collectively designated as HTT. Although all Drosophila telomeres terminate with HTT arrays and are capped by the terminin complex, they differ in the type of subtelomeric chromatin. The HTT sequences of YS, YL, XR, and 4L are juxtaposed to constitutive heterochromatin, while the HTTs of the other telomeres are linked to either the TAS repeat-associated chromatin (XL, 2L, 2R, 3L, 3R) or to the specialized 4R chromatin. We found that mutations in pendolino (peo) cause (telomeric fusions) that preferentially involve the heterochromatin-associated telomeres (Ha-telomeres), a telomeric fusion pattern never observed in the other 10 telomere-capping mutants characterized so far. Peo, is homologous to the E2 variant ubiquitin-conjugating enzymes and is required for DNA replication. Our analyses lead us to hypothesize that DNA replication in Peo-depleted cells results in specific fusigenic lesions concentrated in Ha-telomeres. These data provide the first demonstration that subtelomeres can affect telomere fusion

Whole-mount Senescence-Associated Beta-Galactosidase (SA-β-GAL) Activity Detection Protocol for Adult Zebrafish

International audienceSenescence-associated beta-galactosidase (SA-β-GAL) is an enzyme that accumulates in the lysosomes of senescent cells, where it hydrolyses β-galactosides. With p16, it represents a well-recognized biomarker used to assess senescence both in vivo and in cell culture. The use of a chromogenic substrate, such as 5-bromo-4-chloro-3-indoylβ-d-galactopyranoside (X-Gal), allows the detection of SA-β-GAL activity at pH 6.0 by the release of a visible blue product. Senescence occurs during aging and is part of the aging process itself. We have shown that prematurely aged zebrafish accumulate senescent cells detectable by SA-β-GAL staining in different tissues, including testis and gut. Here, we report a detailed protocol to perform an SA-β-GAL assay to detect senescent cell accumulation across the entire adult zebrafish organism (Danio rerio). We also identify previously unreported organs that show increased cell senescence in telomerase mutants, including the liver and the spinal cord. Cite as: Marzullo, M. et al. (2022). Whole-mount Senescence-Associated Beta-Galactosidase (SA-β-GAL) Activity Detection Protocol for Adult Zebrafish. Bio-protocol 12(13): e4457

Modeling Myotonic Dystrophy Type 2 Using <i>Drosophila melanogaster</i>

Myotonic dystrophy 2 (DM2) is a genetic multi-systemic disease primarily affecting skeletal muscle. It is caused by CCTGn expansion in intron 1 of the CNBP gene, which encodes a zinc finger protein. DM2 disease has been successfully modeled in Drosophila melanogaster, allowing the identification and validation of new pathogenic mechanisms and potential therapeutic strategies. Here, we describe the principal tools used in Drosophila to study and dissect molecular pathways related to muscular dystrophies and summarize the main findings in DM2 pathogenesis based on DM2 Drosophila models. We also illustrate how Drosophila may be successfully used to generate a tractable animal model to identify novel genes able to affect and/or modify the pathogenic pathway and to discover new potential drugs

Su(var)3-9 mediates age-dependent increase in H3K9 methylation on TDP-43 promoter triggering neurodegeneration

Abstract Aging progressively modifies the physiological balance of the organism increasing susceptibility to both genetic and sporadic neurodegenerative diseases. These changes include epigenetic chromatin remodeling events that may modify the transcription levels of disease-causing genes affecting neuronal survival. However, how these events interconnect is not well understood. Here, we found that Su(var)3-9 causes increased methylation of histone H3K9 in the promoter region of TDP-43, the most frequently altered factor in amyotrophic lateral sclerosis (ALS), affecting the mRNA and protein expression levels of this gene through epigenetic modifications that appear to be conserved in aged Drosophila brains, mouse, and human cells. Remarkably, augmented Su(var)3-9 activity causes a decrease in TDP-43 expression followed by early defects in locomotor activities. In contrast, decreasing Su(var)3-9 action promotes higher levels of TDP-43 expression, improving motility parameters in old flies. The data uncover a novel role of this enzyme in regulating TDP-43 expression and locomotor senescence and indicate conserved epigenetic mechanisms that may play a role in the pathogenesis of ALS

Opposing p53 and mTOR/AKT promote an in vivo switch from apoptosis to senescence upon telomere shortening in zebrafish

Progressive telomere shortening during lifespan is associated with restriction of cell proliferation, genome instability and aging. Apoptosis and senescence are the two major outcomes upon irreversible cellular damage. Here, we show a transition of these two cell fates during aging of telomerase deficient zebrafish. In young telomerase mutants, proliferative tissues exhibit DNA damage and p53-dependent apoptosis, but no senescence. However, these tissues in older animals display loss of cellularity and senescence becomes predominant. Tissue alterations are accompanied by a pro-proliferative stimulus mediated by AKT signaling. Upon AKT activation, FoxO transcription factors are phosphorylated and translocated out of the nucleus. This results in reduced SOD2 expression causing an increase of ROS and mitochondrial dysfunction. These alterations induce p15/16 growth arrest and senescence. We propose that, upon telomere shortening, early apoptosis leads to cell depletion and insufficient compensatory proliferation. Following tissue damage, the mTOR/AKT is activated causing mitochondrial dysfunction and p15/ 16-dependent senescence