Organismal aging is influenced by a multitude of intrinsic and extrinsic factors, and heterochromatin loss has been proposed to be one of the causes of aging. However, the role of heterochromatin in animal aging has been controversial. Here we show that heterochromatin formation prolongs lifespan and controls ribosomal RNA synthesis in Drosophila. Animals with decreased heterochromatin levels exhibit a dramatic shortening of lifespan, whereas increasing heterochromatin prolongs lifespan. The changes in lifespan are associated with changes in muscle integrity. Furthermore, we show that heterochromatin levels decrease with normal aging and that heterochromatin formation is essential for silencing rRNA transcription. Loss of epigenetic silencing and loss of stability of the rDNA locus have previously been implicated in aging of yeast. Taken together, these results suggest that epigenetic preservation of genome stability, especially at the rDNA locus, and repression of unnecessary rRNA synthesis, might be an evolutionarily conserved mechanism for prolonging lifespan.

STAT (Signal transducer and activator of transcription) is a potent transcription factor and its aberrant activation by phosphorylation is associated with human cancers [1][2][3][4] . We have shown previously that overactivation of JAK, which phosphorylates STAT 5,6 , disrupts heterochromatin formation globally in Drosophila melanogaster 7 . However, it remains unclear how this effect is mediated and whether STAT is involved. Here, we demonstrate that Drosophila STAT (STAT92E) is involved in controlling heterochromatin protein 1 (HP1) distribution and heterochromatin stability. We found, unexpectedly, that loss of STAT92E, had the same effects as overactivation of JAK in disrupting heterochromatin formation and heterochromatic gene silencing, whereas overexpression of STAT92E had the opposite effects. We have further shown that the unphosphorylated or 'transcriptionally inactive' form of STAT92E is localized on heterochromatin in association with HP1, and is required for stabilizing HP1 localization and histone H3 Lys 9 methylation (H3mK9). However, activation by phosphorylation reduces heterochromatin-associated STAT92E, causing HP1 displacement and heterochromatin destabilization. Thus, reducing levels of unphosphorylated STAT92E, either by loss of STAT92E or increased phosphorylation, causes heterochromatin instability. These results suggest that activation of STAT by phosphorylation controls both access to chromatin and activity of the transcription machinery.To understand the molecular mechanism underlying JAK/STAT-mediated tumour formation, we have previously investigated the role of JAK in a Drosophila leukaemia model, in which a hyperactive mutant form of JAK (Tum-1) causes leukaemia-like overproliferation of blood cells 5,8 . We have demonstrated that oncogenic JAK disrupts heterochromatin formation globally, allowing transcriptional activation of genes that are not necessarily direct targets of STAT 7,9 . The molecular mechanism underlying the effects of Hopscotch (Hop, Drosophila JAK) on heterochromatin remains unclear. It may be mediated by phosphorylation of STAT92E, as in the canonical JAK/STAT pathway 10,11 . Alternatively, Hop may activate cellular targets other than STAT92E 9 .To investigate whether disruption of heterochromatin induced by Hop-activation 7 is mediated by STAT92E, we examined the effects of reducing stat92E + dosage on heterochromatic gene silencing, which can be measured by position-effect variegation (PEV) 12 We next examined the epistatic relationship between HP1 and STAT92E ( Fig. 1d-k). HP1, encoded by Su(var)205, is a constitutive component of heterochromatin and is essential for heterochromatic gene silencing 12,13 . Consistent with the results described above, we found that increasing stat92E + dosage by a chromosomal duplication (referred to as 3 × stat92E + , Fig. 1f) or a stat92E + transgene ( Supplementary Information, Fig. S1) enhanced heterochromatic gene silencing, resulting in complete silencing of the variegated white + gene. This effect was antagonized,...

Safety concerns have been raised over the extensive applications of silver nanoparticles (AgNPs) because nano dimensions make them highly bioactive, being potentially harmful to the exposed humans. Surface physico-chemistry (shape, surface charge, chemical composition, etc.) that mainly dictates nano-bio interactions is relevant for influencing their biocompatibility and toxicity. Although the hazardousness of AgNPs has been demonstrated in vitro and in vivo, mechanistic understanding of the toxicity particularly at the molecular and organismal levels, in addition to oxidative stress and silver ion dissolution, has remained unclear. A growing body of research has elucidated that autophagy, being activated in response to exposure to various nanomaterials, may serve as a cellular defense mechanism against nanotoxicity. Recently, autophagy activation was shown to correlate with AgNPs exposure; however, the subsequent autophagosome-lysosome fusion was defective. As autophagy plays a crucial role in selective removal of stress-mediated protein aggregates and injured organelles, AgNPs-induced autophagic flux defect may consequently lead to aggravated cytotoxic responses. Furthermore, we suggest that p62 accumulation resulting from defective autophagy may also potentially account for AgNPs cytotoxicity. Intriguingly, AgNPs have been shown to interfere with ubiquitin modifications, either via upregulating levels of enzymes participating in ubiquitination, or through impairing the biological reactivity of ubiquitin (due to formation of AgNPs-ubiquitin corona). Ubiquitination both confers selectivity to autophagy as well as modulates stabilization, activation, and trafficking of proteins involved in autophagic clearance pathways. In this regard, we offer a new perspective that interference of AgNPs with ubiquitination may account for AgNPs-induced defective autophagy and cytotoxic effects.

Silver nanoparticles (AgNPs) are widely used in the household, medical and industrial sectors due to their effective bactericidal activities and unique plasmonic properties. Despite the promising advantages, safety concerns have been raised over the usage of AgNPs because they pose potential hazards. However, the mechanistic basis behind AgNPs toxicity, particularly the sublethal effects at the organismal level, has remained unclear. In this study, we used a powerful in vivo platform Drosophila melanogaster to explore a wide spectrum of adverse effects exerted by dietary AgNPs at the organismal, cellular and molecular levels. Lethal doses of dietary AgNPs caused developmental delays and profound lethality in developing animals and young adults. In contrast, exposure to sublethal doses, while not deadly to developing animals, shortened the adult lifespan and compromised their tolerance to oxidative stress. Importantly, AgNPs mechanistically resulted in tissue-wide accumulation of reactive oxygen species (ROS) and activated the Nrf2-dependent antioxidant pathway, as demonstrated by an Nrf2 activity reporter in vivo. Finally, dietary AgNPs caused a variety of ROS-mediated stress responses, including apoptosis, DNA damage, and autophagy. Altogether, our study suggests that lethal and sublethal doses of AgNPs, have acute and chronic effects, respectively, on development and longevity by inducing ROS-mediated stress responses.

Heterochromatin is a form of highly compacted chromatin associated with epigenetic gene silencing and chromosome organization. We have previously shown that unphosphorylated nuclear signal transducer and activator of transcription (STAT) physically interacts with heterochromatin protein 1 (HP1) to promote heterochromatin stability. To understand whether STAT and heterochromatin are important for maintenance of genome stability, we genetically manipulated the levels of unphosphorylated STAT and HP1 [encoded by Su(var)205] in Drosophila and examined the effects on chromosomal morphology and resistance to DNA damage under conditions of genotoxic stress. Here we show that, compared with wild-type controls, Drosophila mutants with reduced levels of unphosphorylated STAT or heterochromatin are more sensitive to radiation-induced cell cycle arrest, have higher levels of spontaneous and radiation-induced DNA damage, and exhibit defects in chromosomal compaction and segregation during mitosis. Conversely, animals with increased levels of heterochromatin exhibit less DNA damage and increased survival rate after irradiation. These results suggest that maintaining genome stability by heterochromatin formation and correct chromosomal packaging is essential for normal cellular functions and for survival of animals under genotoxic stress.

The dynamic regulation of chromatin structure by histone post-translational modification is an essential regulatory mechanism that controls global gene transcription. The Kdm4 family of H3K9me2,3 and H3K36me2,3 dual specific histone demethylases has been implicated in development and tumorigenesis. Here we show that Drosophila Kdm4A and Kdm4B are together essential for mediating ecdysteroid hormone signaling during larval development. Loss of Kdm4 genes leads to globally elevated levels of the heterochromatin marker H3K9me2,3 and impedes transcriptional activation of ecdysone response genes, resulting in developmental arrest. We further show that Kdm4A interacts with the Ecdysone Receptor (EcR) and colocalizes with EcR at its target gene promoter. Our studies suggest that Kdm4A may function as a transcriptional co-activator by removing the repressive histone mark H3K9me2,3 from cognate promoters.

We have previously reported the isolation of the JAK1 gene from the round-spotted pufferfish. In the present study, we cloned and characterized genomic sequences encoding pufferfish JAK2, JAK3, and TYK2, which are other members of JAK family. To our knowledge, this is the first report to demonstrate the existence of four JAK genes in fish. All pufferfish JAK genes except JAK1 are composed of 24 exons; JAK1 has an additional exon. A comparison of the exon-intron organization of these genes revealed that the splice sites of JAK genes are nearly identical. In addition, all pufferfish JAK genes have one intron in the 5' untranslated region. Taken together, these data suggest that the pufferfish JAK genes may have evolved from a common ancestor. By 5' rapid amplification of cDNA ends and sequence analysis, we deduced the promoter regions for all JAK genes and found they do not contain typical TATA or CCAAT boxes but rather numerous other potential binding sites for transcription factors. Interestingly, the TYK2 gene is linked to CDC37 in a head-to-tail manner with a small intergenic region of 292 bp. Within this region, there are two potential binding sites for transcriptional factors such as c-Myb and NF-IL6. The putative promoter regions of all JAK genes were tested either in a carp CF cell line or in zebrafish embryos using CAT or lacZ as reporter genes. Both assays confirmed the transcriptional activities of these promoters in vitro and in vivo.

Both epidemiologic and laboratory studies have shown the chemopreventive effects of 1a, 25-dihydroxyvitamin D 3 (1,25-VD) in tumorigenesis. However, understanding of the molecular mechanism by which 1,25-VD prevents tumorigenesis remains incomplete. In this study, we used an established mouse model of chemical carcinogenesis to investigate how 1,25-VD prevents malignant transformation. In this model, 1,25-VD promoted expression of the DNA repair genes RAD50 and ATM, both of which are critical for mediating the signaling responses to DNA damage. Correspondingly, 1,25-VD protected cells from genotoxic stress and growth inhibition by promoting double-strand break DNA repair. Depletion of the vitamin D receptor (VDR) reduced these genoprotective effects and drove malignant transformation that could not be prevented by 1,25-VD, defining an essential role for VDR in mediating the anticancer effects of 1,25-VD. Notably, genotoxic stress activated ATM and VDR through phosphorylation of VDR. Mutations in VDR at putative ATM phosphorylation sites impaired the ability of ATM to enhance VDR transactivation activity, diminishing 1,25-VD-mediated induction of ATM and RAD50 expression. Together, our findings identify a novel vitamin Dmediated chemopreventive mechanism involving a positive feedback loop between the DNA repair proteins ATM and VDR. Cancer Res; 72(4); 958-68. Ó2011 AACR.