Oncogene-induced autophagy and the Goldilocks principle

Although several oncogenes enhance autophagic flux, the molecular mechanism and consequences of oncogene-induced autophagy remain to be clarified. We have recently shown that expression of oncogenic H-Ras (V12) promotes autophagy through upregulation of Beclin 1 and the BH3-only protein Noxa. H-Ras-expressing cells undergo autophagic cell death as a result of Noxa-mediated displacement of Mcl-1 and Bcl-xL from Beclin 1. Oncogenic H-Ras-induced death is attenuated through knockdown of BECLIN 1, ATG5, or ATG7, or through overexpression of Mcl-1, Bcl-2, Bcl-xL and their close relatives. These observations suggest that high-intensity oncogene activation may be selected against by promoting excessive autophagy, leading to cell death. Consequently, such oncogenes may select for cells with a reduced capacity for autophagy, either through loss of a BECLIN 1 allele or through upregulation of negative regulators of Beclin 1, such as Bcl-2 family members.     

Jailbreak: Oncogene-induced senescence and its evasion

Aberrant oncogenic signals are typically counteracted by anti-proliferative mechanisms governed principally by the p53 and Rb tumour-suppressor proteins. Apoptosis is firmly established as a potent anti-proliferative mechanism to prevent tumour growth but it is only in recent years that oncogene-induced senescence has achieved similar recognition. Senescence is defined as an irreversible cell-cycle arrest suggesting that entry of oncogene-expressing cells into this static yet viable state is permanent. However, tumours do develop and express the very same oncogenes that landed them in jail. We ask whether this is because rogue incipient cancer cells find ways to escape this imposed imprisonment or otherwise entirely avoid capture by senescence gate-keepers.     

Goldilocks,vitamin D and sarcoidosis

While low levels of vitamin D can increase the risk for osteoporosis, excessive amounts of vitamin D may also be problematic. Hypercalcemia and hypercalcuria due to increased vitamin D activity occur in a significant proportion of sarcoidosis patients. Saidenberg-Kermanac’h and colleagues compared vitamin D levels with bone fragility fractures in their sarcoidosis clinic. They found that a 25-(OH) vitamin D level between 10 and 20 ng/ml was associated with the lowest risk of bone fractures and paradoxically higher levels increased the risk of bone fractures. Using less vitamin D supplementation may simultaneously lower the risk for bone fracture and hypercalcemia in sarcoidosis.In the previous issue, Saidenberg-Kermanac’h and colleagues provide more information regarding the complexity of vitamin D activity in sarcoidosis [1]. A few years ago, vitamin D was declared the nutrient of the decade. This was heady stuff for a sterol that was originally felt important only in preventing rickets. Studies have demonstrated its key role in calcium absorption and bone growth. Beyond that, vitamin D has been considered an important sterol in various aspects of health. Low levels of vitamin D have been associated with increased risk for cancer, type 2 diabetes, and heart disease.Most of these observations have been based on measurements of 25-(OH)-vitamin D3 (ergocalcitrol). This sterol is converted by 1-alpha hydroxylase to 1,25-(OH)2-vitamin D3 (calcitrol), the active form of vitamin D. This conversion occurs in the kidney and patients with chronic renal failure require calcitrol replacement.In tuberculosis, vitamin D supplementation has been recommended in patients because vitamin D is crucial in the granulomatous reaction against the organism. However, what may be good for tuberculosis eradication may not be good for sarcoidosis. It has been noted that excessive amounts of vitamin D are associated with a worse clinical outcome in sarcoidosis [2]. In granulomas, there may be increased activity of 1-alpha hydroxylase. As part of the Th-1 immune response, calcitrol has a paracrine effect within the granuloma. In some cases, this leads to excessive calcitrol, resulting in hypercalcuria or hypercalcemia [3]. At least 10% of sarcoidosis patients have hypercalcemia, half of whom can develop associated renal dysfunction [3,4]. In some cases, hypercalcemic renal failure can be reversed by simply withdrawing vitamin D supplementation [3]. There are case reports of excessive vitamin D replacement leading to hypercalcemia in patients with mycobacterial infections [5].The sarcoidosis patient may be treated with glucocorticoids, sometimes for years. Obviously, long-term glucocorticoid administration places the patient at risk for developing osteoporosis [6,7]. In rheumatoid arthritis, patients undergoing prolonged glucocorticoid treatment are recommended to receive calcium and vitamin D replacement [8]. While this is the cornerstone of prevention of osteoporosis, the role of calcium and vitamin D replacement in sarcoidosis remains unclear [9].Into this quandary comes the study by Saidenberg-Kermanac’h and colleagues reported in the previous issue of Arthritis Research & Therapy[1]. After studying a large cohort of sarcoidosis patients from their clinic, the authors found that fragility fractures occurred in nearly a quarter of them. The fracture risk was increased for those treated with corticosteroids. Although low levels of ergocalcitrol was an independent risk for osteoporosis, ironically high levels of ergocalcitrol were also associated with an increased risk for osteoporosis. They found that ergocalcitrol levels of 10 to 20 ng/ml was associated with the lowest fracture risk for patients. This J shaped risk for bone fragility has been noted in non-sarcoidosis patients, although the proposed target levels are higher for these patients [10]. For the clinician treating sarcoidosis, one has to balance not only the risk for osteoporosis, but also the risk for hypercalcemia and renal failure (Figure 1).Open in a separate windowFigure 1Schematic depicting vitamin D metabolism in the body from ergocalcitrol to calcitrol. The conversion is enhanced by increased activity of 1-alpha hydroxylase activity in the granuloma of sarcoidosis patients. The untoward consequences of low or high vitamin D activity are summarized at the bottom of the figure.One possible explanation for the lower ideal ergocalcitrol level in sarcoidosis is the enhanced activity of 1-alpha hydroxylase in sarcoidosis granulomas. The authors did not provide information regarding calcitrol levels in their patients. The proportion of calcitrol to ergocalcitrol appears to be higher in sarcoidosis compared to non-sarcoidosis conditions. In one study of 270 sarcoidosis patients, 80% had low ergocalcitrol levels, but less than 1% had low calcitrol levels. In fact, that study found that 10% of patients had elevated calcitrol levels [3]. Those with elevated calcitrol were more likely to have a history of hypercalcemia or hypercalcuria. Higher levels of calcitrol have been associated with more advanced pulmonary sarcoidosis [2].The other potential benefits of vitamin D replacement in sarcoidosis are unclear. Should sarcoidosis patients with low ergocalcitrol but normal calcitrol levels be prescribed vitamin D supplementation to reduce their risk for cancer and type 2 diabetes? If so, do they increase their risk for hypercalcemia or hypercalcuria? Could this increased vitamin D intake raise the functional level of vitamin D even higher and therefore increase the risk for osteoporosis?To paraphrase Goldilocks, one does not want too little or too much vitamin D. You want just the right amount.     

Oncogene-induced senescence pathways weave an intricate tapestry

The induction of cellular senescence by activated oncogenes acts as a barrier to cell transformation. Now, identify a key component of a senescence pathway that prevents tumorigenesis in a mouse model of skin cancer. They show that the p38-regulated/activated protein kinase (PRAK) induces senescence downstream of oncogenic Ras by directly phosphorylating and activating the tumor-suppressor protein p53.     

Oncogene-induced senescence results in marked metabolic and bioenergetic alterations

Oncogene-induced senescence (OIS) is characterized by permanent growth arrest and the acquisition of a secretory, pro-inflammatory state. Increasingly, OIS is viewed as an important barrier to tumorgenesis. Surprisingly, relatively little is known about the metabolic changes that accompany and therefore may contribute to OIS. Here, we have performed a metabolomic and bioenergetic analysis of Ras-induced senescence. Profiling approximately 300 different intracellular metabolites reveals that cells that have undergone OIS develop a unique metabolic signature that differs markedly from cells undergoing replicative senescence. A number of lipid metabolites appear uniquely increased in OIS cells, including a marked increase in the level of certain intracellular long chain fatty acids. Functional studies reveal that this alteration in the metabolome reflects substantial changes in overall lipid metabolism. In particular, Ras-induced senescent cells manifest a decline in lipid synthesis and a significant increase in fatty acid oxidation. Increased fatty acid oxidation results in an unexpectedly high rate of basal oxygen consumption in cells that have undergone OIS. Pharmacological or genetic inhibition of carnitine palmitoyltransferase 1, the rate-limiting step in mitochondrial fatty acid oxidation, restores a pre-senescent metabolic rate and, surprisingly, selectively inhibits the secretory, pro-inflammatory state that accompanies OIS. Thus, Ras-induced senescent cells demonstrate profound alterations in their metabolic and bioenergetic profiles, particularly with regards to the levels, synthesis and oxidation of free fatty acids. Furthermore, the inflammatory phenotype that accompanies OIS appears to be related to these underlying changes in cellular metabolism.     

Oncogene-induced senescence results in marked metabolic and bioenergetic alterations

Oncogene-induced senescence (OIS) is characterized by permanent growth arrest and the acquisition of a secretory, pro-inflammatory state. Increasingly, OIS is viewed as an important barrier to tumorgenesis. Surprisingly, relatively little is known about the metabolic changes that accompany and therefore may contribute to OIS. Here, we have performed a metabolomic and bioenergetic analysis of Ras-induced senescence. Profiling approximately 300 different intracellular metabolites reveals that cells that have undergone OIS develop a unique metabolic signature that differs markedly from cells undergoing replicative senescence. A number of lipid metabolites appear uniquely increased in OIS cells, including a marked increase in the level of certain intracellular long chain fatty acids. Functional studies reveal that this alteration in the metabolome reflects substantial changes in overall lipid metabolism. In particular, Ras-induced senescent cells manifest a decline in lipid synthesis and a significant increase in fatty acid oxidation. Increased fatty acid oxidation results in an unexpectedly high rate of basal oxygen consumption in cells that have undergone OIS. Pharmacological or genetic inhibition of carnitine palmitoyltransferase 1, the rate-limiting step in mitochondrial fatty acid oxidation, restores a presenescent metabolic rate and, surprisingly, selectively inhibits the secretory, pro-inflammatory state that accompanies OIS. Thus, Ras-induced senescent cells demonstrate profound alterations in their metabolic and bioenergetic profiles, particularly with regards to the levels, synthesis and oxidation of free fatty acids. Furthermore, the inflammatory phenotype that accompanies OIS appears to be related to these underlying changes in cellular metabolism.Key words: oncogene-induced senescence, metabolomics, Ras, fatty acid oxidation     

Oncogene-induced senescence pathways in melanocytes

Comment on: Haferkamp S, et al. Aging 2009; 1:542-56.     

Oncogene-induced reactive oxygen species fuel hyperproliferation and DNA damage response activation

Oncogene-induced reactive oxygen species (ROS) have been proposed to be signaling molecules that mediate proliferative cues. However, ROS may also cause DNA damage and proliferative arrest. How these apparently opposite roles can be reconciled, especially in the context of oncogene-induced cellular senescence, which is associated both with aberrant mitogenic signaling and DNA damage response (DDR)-mediated arrest, is unclear. Here, we show that ROS are indeed mitogenic signaling molecules that fuel oncogene-driven aberrant cell proliferation. However, by their very same ability to mediate cell hyperproliferation, ROS eventually cause DDR activation. We also show that oncogenic Ras-induced ROS are produced in a Rac1 and NADPH oxidase (Nox4)-dependent manner. In addition, we show that Ras-induced ROS can be detected and modulated in a living transparent animal: the zebrafish. Finally, in cancer we show that Nox4 is increased in both human tumors and a mouse model of pancreatic cancer and specific Nox4 small-molecule inhibitors act synergistically with existing chemotherapic agents.     

Goldilocks mushrooms: How ballistospory has shaped basidiomycete evolution

Ballistospory has been a governing factor in mushroom diversification. Modifications to fruit body morphology are subject to a series of fundamental constraints imposed by this uniquely fungal mechanism. Gill spacing in lamellate mushrooms, tube width in poroid species, and other configurations of the hymenium must comply with the distance that spores shoot themselves from their basidia. This reciprocal relationship between the development of fruit bodies and spores may have been maintained by a form of evolutionary seesaw proposed in this article. The necessity of the accurate gravitropic orientation of gills and tubes is another constraint on mushroom development and physiology, along with the importance of evaporative cooling of the hymenium for successful spore discharge and the aerodynamic shaping of the fruit body to aid dispersal. Ballistospory has been lost in secotioid and gasteroid basidiomycetes whose spores are dispersed by animal vectors and has been replaced by alterative mechanisms of active spore discharge in some species. Partnered with the conclusions drawn from molecular phylogenetic research, the biomechanical themes discussed in this review afford new ways to think about the evolution of basidiomycetes.     

Oncogene-induced telomere dysfunction enforces cellular senescence in human cancer precursor lesions

In normal human somatic cells, telomere dysfunction causes cellular senescence, a stable proliferative arrest with tumour suppressing properties. Whether telomere dysfunction-induced senescence (TDIS) suppresses cancer growth in humans, however, is unknown. Here, we demonstrate that multiple and distinct human cancer precursor lesions, but not corresponding malignant cancers, are comprised of cells that display hallmarks of TDIS. Furthermore, we demonstrate that oncogenic signalling, frequently associated with initiating cancer growth in humans, dramatically affected telomere structure and function by causing telomeric replication stress, rapid and stochastic telomere attrition, and consequently telomere dysfunction in cells that lack hTERT activity. DNA replication stress induced by drugs also resulted in telomere dysfunction and cellular senescence in normal human cells, demonstrating that telomeric repeats indeed are hypersensitive to DNA replication stress. Our data reveal that TDIS, accelerated by oncogene-induced DNA replication stress, is a biological response of cells in human cancer precursor lesions and provide strong evidence that TDIS is a critical tumour suppressing mechanism in humans.     

Shigella and autophagy

Bacterial invasion of eukaryotic cells, and host recognition and elimination of the invading bacteria, determines the fate of bacterial infection. Once inside mammalian cells, many pathogenic bacteria enter the host cytosol to escape from the lytic compartment and gain a replicative niche. Recent studies indicate that autophagy also recognizes intracellular bacteria. Although autophagy is a conserved membrane trafficking pathway in eukaryotic cells that sequesters undesirable or recyclable cytoplasmic components or organelles and delivers them to lysosomes, autophagy has recently been described as playing a pivotal role as an intracellular surveillance system for recognition and eradication of the pathogens that have invaded the cytoplasm. Indeed, unless they are able to circumvent entrapping by autophagosomes, bacteria ultimately undergo degradation by delivery into autolysosomes. In this review we discuss recent discoveries regarding Shigella strategies for infecting mammalian cells, and then focus on recent studies of an elegant bacterial survival strategy against autophagic degradation.     

Zinc and autophagy

Autophagy is a highly conserved degradative process through which cells overcome stressful conditions. Inasmuch as faulty autophagy has been associated with aging, neuronal degeneration disorders, diabetes, and fatty liver, autophagy is regarded as a potential therapeutic target. This review summarizes the present state of knowledge concerning the role of zinc in the regulation of autophagy, the role of autophagy in zinc metabolism, and the potential role of autophagy as a mediator of the protective effects of zinc. Data from in vitro studies consistently support the notion that zinc is critical for early and late autophagy. Studies have shown inhibition of early and late autophagy in cells cultured in medium treated with zinc chelators. Conversely, excess zinc added to the medium has shown to potentiate the stimulation of autophagy by tamoxifen, H₂O₂, ethanol and dopamine. The potential role of autophagy in zinc homeostasis has just begun to be investigated. Increasing evidence indicates that autophagy dysregulation causes significant changes in cellular zinc homeostasis. Autophagy may mediate the protective effect of zinc against lipid accumulation, apoptosis and inflammation by promoting degradation of lipid droplets, inflammasomes, p62/SQSTM1 and damaged mitochondria. Studies with humans and animal models are necessary to determine whether autophagy is influenced by zinc intake.     

Senescence-associated Long Non-coding RNA (SALNR) Delays Oncogene-induced Senescence through NF90 Regulation

Long non-coding RNAs (lncRNAs) have recently emerged as key players in many physiologic and pathologic processes. Although many lncRNAs have been identified, few lncRNAs have been characterized functionally in aging. In this study, we used human fibroblast cells to investigate genome-wide lncRNA expression during cellular senescence. We identified 968 down-regulated lncRNAs and 899 up-regulated lncRNAs in senescent cells compared with young cells. Among these lncRNAs, we characterized a senescence-associated lncRNA (SALNR), whose expression was reduced during cellular senescence and in premalignant colon adenomas. Overexpression of SALNR delayed cellular senescence in fibroblast cells. Furthermore, we found that SALNR interacts with NF90 (nuclear factor of activated T-cells, 90 kDa), an RNA-binding protein suppressing miRNA biogenesis. We demonstrated that NF90 is a SALNR downstream target, whose inhibition led to premature senescence and enhanced expressions of senescence-associated miRNAs. Moreover, our data showed that Ras-induced stress promotes NF90 nucleolus translocation and suppresses its ability to suppress senescence-associated miRNA biogenesis, which could be rescued by SALNR overexpression. These data suggest that lncRNA SALNR modulates cellular senescence at least partly through changing NF90 activity.