GECKIL LAB:Cutting-edge science

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• Cancer prevention through a diet that affects your epigenome

Cancer, the leading cause of death worldwide, caused 8.2 million deaths in 2012. With 575,000 deaths attributable to cancer in 2010 in the United States, cancer-related deaths in the US are second only to those caused by heart disease, which caused 594,000. How can we end cancer? First and foremost, focus on prevention—the most viable option as a cure.

Historically, cancer has been perceived as a disease in which our genetic makeup dictates our likelihood of developing cancer. Presently, it has become broadly recognized that the initiation and progression of cancer is an intricate web of both genetic makeup and epigenetic events that alter our gene expression. Many studies have proven that epigenetic alterations are key components of the initiation and progression of cancer. These epigenetic processes—including DNA methylation, histone modification, and microRNA expression—are potentially reversible.

CpG island hypermethylation and down-regulation, histon acetylation and the resulting up-regulation of genes are common for many genes involved in a broad range of functions that are deregulated in cancer.

Dietary compounds have been shown to elicit epigenetic changes in cancer cells. To fully understand how we can modulate cancer prevention through lifestyle, research must focus on how diet and bioactive food components specifically impact epigenetic processes. Antioxidants such as carotenoids and fiber found in many vegetables and fruit offer a variety of anti-cancer benefits. Increased dietary folate, a soluble form of B6 vitamin, consumption has been linked to a decrease in colorectal cancer through its affect on DNA methylation. Dietary phytochemicals, that act as anti-cancer agents (including polyphenols, genistein, sulforaphane, resveratrol, and curcumin) have been shown to act through epigenetic mechanisms.

Cancer prevention is the best way to ultimately cure the disease. To work towards cancer prevention, we must further explore how dietary modifications may achieve epigenetic reprogramming, resulting in the maintenance of normal gene expression and reversal of tumor progression. (Scizzle, FEBRUARY 4, 2014 • KELLY JAMIESON THOMAS)

• Healing cancer cells and aerobic glycolysis

Instead of relying on mitochondrial oxidative phosphorylation, most cancer cells rely heavily on aerobic glycolysis, a phenomenon termed as “the Warburg effect. This effect may be is a direct consequence of damage and it persists in cancer cells that recover from damage. Glycolysis and rate of cell proliferation in cancer cells that recovered from severe damage show that such in vitro Damage-Recovered (DR) cells exhibit mitochondrial structural remodeling, display Warburg effect, and show increased in vitro and in vivo proliferation and tolerance to damage.

To test whether cancer cells derived from tumor microenvironment can show similar properties, (DR) cells from tumors show increased aerobic glycolysis and a high growth rate. These findings show that Warburg effect and its consequences are induced in cancer cells that survive severe damage. (Biochemical and biophysical research communications. 2014 May 24, PMID: 24802411).

• Lower energy flux and higher aerobic glycolysis: constant growth

Fermentating glucose in the presence of enough oxygen to support respiration, known as aerobic glycolysis (a.k.a. Warburg effect) is believed to maximize growth rate. The cells support a constant biomass-production rate with decreasing rates of respiration and ATP production but also decrease their stress resistance. As the respiration rate decreases, so do the levels of enzymes catalyzing rate-determining reactions of the tricarboxylic-acid cycle (providing NADH for respiration) and of mitochondrial folate-mediated NADPH production (required for oxidative defense).

The findings demonstrate that exponential growth can represent not a single metabolic/physiological state but a continuum of changing states and that aerobic glycolysis can reduce the energy demands associated with respiratory metabolism and stress survival. (Cell reports. 2014 Apr 24. PMID: 24767987)

• Stem cells on a diet

Knowing how an organism’s tissues handle stress throughout life is key to understanding ageing and disease. Stems cells of the blood system seem to tackle metabolic stress by means of a process called autophagy.

Stem cells in adult tissues function to replace lost, damaged or diseased cells throughout an organism’s life, thereby helping to maintain tissue health. But what protects the rare, long-lived stem cells from a lifetime of exposure to cellular and environmental stressors such as inflammation, radiation and metabolic alterations?

Aautophagy, a process of cellular self-cannibalization, is one mechanism that haematopoietic stem cells of the blood system use to protect themselves during times of metabolic stress, when nutrients are limited. When cells are starved of nutrients, the tissue’s stem cells must choose whether to live or die. The options are committing suicide by apoptotic cell death, or self-preservation through autophagy, whereby cells recycle damaged or dispensable proteins and organelles into basic components to support cellular growth.

Autophagy is thought to be a major factor in ageing. Loss of autophagy in tissues such as the brain, liver and heart leads to an increase in age-related disorders, including neurodegeneration, metabolic syndromes and cardiac dysfunction. Conversely, factors that stimulate autophagy abrogate these problems and have been linked to greater longevity. It has therefore been hypothesized that reduced autophagy contributes to the diminishing stem-cell function that occurs with age. No study, however, has investigated the direct role of autophagy in adult stem-cell function.

Warr et al. explore this question in both young mouse haematopoietic stem cells (HSCs) and in more-differentiated HSC progeny, including progenitor cells of the immune cells granulocytes and macrophages. The authors find that little or no autophagy occurs in freshly isolated young HSCs, but that this process can be rapidly induced when the cells are exposed to metabolic stress both in vitro and in vivo. Moreover, when autophagy is inhibited during such metabolic stress, young HSCs rapidly die through apoptosis, indicating that autophagy is crucial for their survival. By contrast, granulocyte–macrophage progenitor cells show higher baseline levels of autophagy, but no shift under starvation conditions. Autophagy can be stimulated in several ways, including through inhibition of the signalling molecule mTOR and activation of stress-induced transcription factors such as FoxO3 and p53. Warr and colleagues find that the primary driver in HSCs is FoxO3, with little contribution from mTOR or p53.(Nature 494, 317–318 (21 February 2013) doi:10.1038/nature11948)

• Wnt proteins as morphogens

Wnt signalling molecules are thought to direct the development of an organism by spreading through tissues. But flies grow with almost normal appendages even when their main Wnt protein cannot move.

The Drosophila (fruitfly) protein Wingless (Wg) is the prototype member of the Wnt family of proteins, which regulate tissue patterning and growth during development. Wg is thought to act as a morphogen — a protein that forms concentration gradients as it spreads from its site of synthesis and that regulates gene expression as a function of its concentration.

Wing formation in flies expressing a form of Wg that is tethered to the cell membrane, in place of the secreted protein. Normal wing morphology, although development is delayed and the final wings are smaller than those of normal flies. Morphogen regulation of target genes depends on the physical distance from the morphogen-secreting cell population, such that the levels of this molecule provide a genetic reading of position, a key issue in morphogenesis. The best examples of morphogens come from Drosophila: the secreted molecules Hedgehog, Decapentaplegic (Dpp) and Wingless (Wg) have been identified as morphogens, and for Dpp and Wg there is compelling evidence that they act at long range. It follows from the very definition of a morphogen that the spread of the molecule is an essential component of its function. (Nature 505, 162–163 (09 January 2014) doi:10.1038/nature12848)

• Mitochondrial form and function

Mitochondria are one of the major ancient endomembrane systems in eukaryotic cells. Owing to their ability to produce ATP through respiration, they became a driving force in evolution. As an essential step in the process of eukaryotic evolution, the size of the mitochondrial chromosome was drastically reduced, and the behaviour of mitochondria within eukaryotic cells radically changed.

Recent advances have revealed how the organelle’s behaviour has evolved to allow the accurate transmission of its genome and to become responsive to the needs of the cell and its own dysfunction.

Mitochondria arose around two billion years ago from the engulfment of an α-proteobacterium by a precursor of the modern eukaryotic cell. Although mitochondria have maintained the double membrane character of their ancestors and the core of ATP production, their overall form and composition have been drastically altered, and they have acquired myriad additional functions within the cell.

As part of the process of acquiring new functions during evolution, most of the genomic material of the α-proteobacterium progenitor was rapidly lost or transferred to the nuclear genome. What remains in human cells is a small, approximately 16 kilobase, circular genome, which is present in cells in a vast excess of copies relative to nuclear chromosomes. The human mitochondrial genome contains genetic coding information for 13 proteins, which are core constituents of the mitochondrial respiratory complexes I–IV that are embedded in the inner membrane.

Functioning together with the Krebs’ cycle in the matrix, the respiratory chain creates an electrochemical gradient through the coupled transfer of electrons to oxygen and the transport of protons from the matrix across the inner membrane into the intermembrane space. The electrochemical gradient powers the terminal complex V of the chain, ATP synthase, which is an ancient rotary turbine machine that catalyses the synthesis of most cellular ATP.

The electrochemical potential is harnessed for additional crucial mitochondrial functions, such as buffering the signalling ion Ca2+ through uptake by a uniporter in the inner membrane. A reduction in the electrochemical potential of mitochondria in cells has evolved as a read-out for mitochondrial functional status, which, as discussed later, creates signals to activate pathways that repair and/or eliminate defective mitochondria. (Nature 505, 335–343 (16 January 2014) doi:10.1038/nature12985)

• Competing endogenous RNAs

Recent reports have described an intricate interplay among diverse RNA species, including protein-coding messenger RNAs and non-coding RNAs such as long non-coding RNAs, pseudogenes and circular RNAs. These RNA transcripts act as competing endogenous RNAs (ceRNAs) or natural microRNA sponges — they communicate with and co-regulate each other by competing for binding to shared microRNAs, a family of small non-coding RNAs that are important post-transcriptional regulators of gene expression.

Understanding this novel RNA crosstalk will lead to significant insight into gene regulatory networks and have implications in human development and disease. Aside from around 21,000 protein-coding genes (less than 2% of the total genome), the human transcriptome includes about 9,000 small RNAs, about 10,000–32,000 long non-coding RNAs (lncRNAs) and around 11,000 pseudogenes.

Non-coding transcripts can generally be divided into two major classes on the basis of their size. Small non-coding RNAs have been relatively well characterized, and include transfer RNAs, which are involved in translation of messenger RNAs; microRNAs (miRNAs) and small-interfering RNAs, which are implicated in post-transcriptional RNA silencing; small nuclear RNAs, which are involved in splicing; small nucleolar RNAs, which are implicated in ribosomal RNA modification; PIWI-interacting RNAs, which are involved in transposon repression; and transcription initiation RNAs, promoter upstream transcripts and promoter-associated small RNAs, which may be involved in transcription regulation. lncRNAs can vary in length from 200 nucleotides to 100 kilobases, and have been implicated in a diverse range of biological processes from pluripotency to immune responses.

Although thousands of lncRNAs have been identified in the past decade (one of the best-studied and most dramatic examples is XIST which can recruit chromatin-modifying complexes to inactivate an entire chromosome during dosage compensation), only a small number have been functionally characterized.

Genome utilization among species is substatntially different (for example, the protein-coding genome constitutes almost the entire genome of unicellular yeast, but only 2% of mammalian genomes). The non-coding transcriptome is often dysregulated in cancer. These observations suggest that the non-coding transcriptome is of crucial importance in determining the greater complexity of higher eukaryotes and in disease pathogenesis. (Nature 505, 344–352 (16 January 2014) doi:10.1038/nature12986)

• A niche for haematopoietic stem cells

Niches are local tissue microenvironments that maintain and regulate stem cells. Haematopoiesis provides a model for understanding mammalian stem cells and their niches, but the haematopoietic stem cell (HSC) niche remains incompletely defined.

Outstanding questions concern the cellular complexity of the niche, the role of the endosteum and functional heterogeneity among perivascular microenvironments. Haematopoietic stem cell (HSC) niches are present in diverse tissues throughout development, beginning in the aorta–gonad–mesonephros (AGM) region and the yolk sac, followed by the placenta, fetal liver, spleen and bone marrow.

Postnatally, the bone marrow is the primary site of HSC maintenance and haematopoiesis, but in response to haematopoietic stress the niche can shift to extramedullary sites. Defining niche components and how they work in concert to regulate haematopoiesis provides the opportunity to improve regeneration following injury or HSC transplantation and to understand how disordered niche function could contribute to disease. (Nature 505, 327–334 (16 January 2014) doi:10.1038/nature12984)

• Cancer predisposition genes

Genes in which germline mutations confer highly or moderately increased risks of cancer are called cancer predisposition genes. More than 100 of these genes have been identified, providing important scientific insights in many areas, particularly the mechanisms of cancer causation. Moreover, clinical utilization of cancer predisposition genes has had a substantial impact on diagnosis, optimized management and prevention of cancer.

The recent transformative advances in DNA sequencing hold the promise of many more cancer predisposition gene discoveries, and greater and broader clinical applications. However, there is also considerable potential for incorrect inferences and inappropriate clinical applications. Realizing the promise of cancer predisposition genes for science and medicine will thus require careful navigation. (Nature 505, 302–308 (16 January 2014) doi:10.1038/nature12981)

• Selectivity of a voltage-gated calcium channel

Voltage-gated calcium (CaV) channels catalyse rapid, highly selective influx of Ca2+ into cells despite a 70-fold higher extracellular concentration of Na+. How CaV channels solve this fundamental biophysical problem remains unclear.

Ca2+ ions flow through CaV channels at a rate of ~1 million ions  per second, yet Na+ conductance is >500-fold lower. Such high-fidelity, high-throughput CaV channel performance is important in regulating intracellular processes such as contraction, secretion, neurotransmission and gene expression in many different cell types. Because the extracellular concentration of Na+ is 70-fold higher than Ca2+, these essential biological functions require CaV channels to be highly selective for Ca2+ in preference to Na+, even though Ca2+ and Na+ have nearly identical diameters (~2 Å). (Nature 505, 56–61 (02 January 2014) doi:10.1038/nature12775)

• DNA repair

UvrD helicase is required for nucleotide excision repair, although its role in this process is not well defined.By inducing backtracking, UvrD exposes DNA lesions shielded by blocked RNA polymerase, allowing nucleotide excision repair enzymes to gain access to sites of damage. UvrD is a bona fide transcription elongation factor that contributes to genomic integrity by resolving conflicts between transcription and DNA repair complexes.

Nucleotide excision repair (NER) is the most versatile and evolutionarily conserved mechanism used by prokaryotic and eukaryotic cells to repair diverse types of DNA lesions. In bacteria, the general NER pathway commences when UvrA and UvrB proteins bind damaged DNA and recruit UvrC to cleave the impaired strand on both sides of the lesion. The resulting oligonucleotide is displaced by UvrD and/or DNA polymerase I, which fills the gap using the complementary strand as a template. (Nature 505, 372–377 (16 January 2014) doi:10.1038/nature12928)

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