Cellular Aging and Apoptosis

Third Public Conference: Cellular Aging and Apoptosis
June 22, 2000

Introduction

In the 1990s, cellular aging and apoptosis gained prominence as two of the most exciting topics in biology. Today, science is on the cusp of appreciating how these processes are involved in human disease. This frontier was the focus of a one-day symposium at Harvard Medical School, where the Giovanni Armenise-Harvard Foundation brought together some of the world’s leading researchers on cellular aging and apoptosis. Held on June 22, 2000, the program drew more than 250 enthusiastic researchers, clinicians, and students from HMS, other Boston-area medical and scientific institutions, and five Italian universities where the Foundation underwrites basic scientific research. The aging and apoptosis symposium capped a week of events that began with the naming of an HMS research building for Count Giovanni Auletta Armenise, whose generosity created the Foundation, and continued with the Foundation’s 4th Annual Scientific Symposium.

Although aging cells stop dividing when senescence sets in, they remain metabolically active for a time. In apoptosis, however, healthy cells abruptly decide to kill themselves. “Although both phenomena have been investigated for years, only now are researchers gaining a molecular and biochemical understanding of how they work,” said program chair Peter M. Howley, Fabian Professor of Pathology and head of the Department of Pathology at HMS. It is increasingly clear that some diseases are caused by the inappropriate engagement of these normal cellular processes. A dramatic example of aging gone wrong is Werner’s disease, a syndrome that prematurely transforms children into wizened, debilitated versions of elderly adults. Apoptosis, on the other hand, has been associated with progressive neurological disorders including Alzheimer’s disease and amyotrophic lateral sclerosis. Cancer researchers like Dr. Howley are also intrigued by new findings about cellular aging and apoptosis, because only cells that escape or overcome these processes turn into cancer cells. Ultimately, scientists hope that a better understanding of these basic events will open the door to novel therapies for diseases now considered difficult, or even impossible, to treat.

Contents of the Public Conference

Cancer, Aging and the Double-edged Sword of Cellular Senescence

Speaker

Judith Campisi, Ph.D.

Head, Center for Research and Education in Aging,

Lawrence Berkeley National Laboratory

The conventional wisdom holds that cancer cells are dangerous because they refuse to become senescent and die on schedule, and that normal cells are good because they turn senescent when they should and don’t hang around to become malignant. As sensible as this sounds, there is mounting evidence that too much senescence may be as bad as too little in terms of promoting tumor growth. Dr. Campisi and her colleagues study the pros and cons of senescence at the Lawrence Berkeley National Laboratory in California, where she is Senior Scientist and Head of the Center for Research and Education in Aging. Earlier, she was on the faculty of the Boston University Medical School. Dr. Campisi received her Ph.D. from the State University of New York at Stony Brook and conducted her postdoctoral research with Arthur Pardee at Harvard Medical School. She has received a number of awards for her work on the relationship between aging and cancer.

Topic

Cancer, Aging and the Double-edged Sword of Cellular Senescence

Epidemiologists and practicing physicians have long noted that cancer rates soar in people over 50, an observation usually attributed to the build-up of deleterious genetic mutations with age. But Dr. Campisi puts at least part of the blame on the accumulation of cells with a senescent phenotype, which hang around in certain tissues long after they’ve undergone changes in morphology, behavior and function. They secrete many different molecules, some of which appear to have a “field effect” that promotes malignant changes in nearby cells.

Support for this idea comes from a series of experiments in which preneoplastic epithelial cells were grown either on a lawn of presenescent stromal cells or one where 10-15% of cells were senescent. Dr. Campisi and her colleagues saw significantly more premalignant changes in cells exposed to senescent neighbors. The investigators obtained similar results in nude mice, where they observed a direct relationship between exposure to senescent cells and the size and number of tumors that developed. In mice, a neoplastic mutation was needed as a starting point for oncogenesis; after that, senescence appeared to drive tumor development.

Dr. Campisi speculates that cellular senescence evolved as a cancer suppression mechanism at a time when the life expectancy for humans was far shorter than it is today. Now that people live so much longer, senescence may be an example of antagonistic pleiotropy: a trait selected to optimize fitness early in life turns out to have unselected deleterious effects later on. Although this may sound like depressing news, Dr. Campisi sees it differently. She believes that additional research will discover small molecules that can counteract damaging secretions from old cells that have overstayed their welcome.

A Link Between Silencing, Metabolism and Aging

Speaker

Leonard P. Guarente, Ph.D.

Professor of Biology

Massachusetts Institute of Technology

Although calorie restriction has repeatedly been shown to increase the longevity of rats, mice, and various other animals, it is not clear why the skinny animals outlive their husky peers. Working in simpler models, including yeast and C. elegans, Dr. Guarente has uncovered a highly conserved gene that may bridge the conceptual gap between energy status and life span. He has carried out this work as a Professor of Biology at the Massachusetts Institute of Technology in Cambridge, having previously been on the Harvard University faculty. Dr. Guarente has a long association with the two institutions, having earned his Ph.D. at Harvard after completing undergraduate work at MIT. His awards and lectureships include election to the American Academy of Microbiology in 1998, and being named Novartis Professor of Biology in 2000. Dr. Guarente is a member of several editorial boards, including Genes and Development, Trends in Genetics, and Journal of Anti-Aging Medicine.

Topic

A Link Between Silencing, Metabolism and Aging

Proposed molecular explanations for aging include genome instability, oxidative damage, and changes in chromatin structure. Of the three, Dr. Guarente’s data indicate that chromatin structure is the most important mechanism governing aging in yeast. Much of his research focuses on SIR2, a key gene for determining whether genetic material is active or silent. SIR2 can repress formation of ribosomal DNA circles that accumulate and are associated with aging in yeast. Yeast cells typically divide 20 to 25 times before entering senescence; in Dr. Guarente’s experiments, deleting SIR2 reduced this number by half, so that yeast cells aged twice as fast. When extra SIR2 was added to the cells, on the other hand, their life span increased. SIR2 encodes a NAD-dependent histone deacetylase, which apparently silences aging-related parts of the yeast genome through selective deacetylation.

Knowing that life extension has often been linked with calorie restriction, Dr. Guarente and his colleagues decided to test this idea in their model. When they limited the glucose available to yeast cells, life span was extended only if both SIR2 and adequate levels of NAD were present. If NAD synthesis was disrupted or SIR2 deleted, the cells lived no longer than amply nourished yeast cells.

Moving up the ladder to C. elegans, Dr. Guarente showed that transgenic worms given extra copies of SIR2 outlived their less well-endowed peers. Higher animals, unlike yeast, don’t accumulate rDNA circles with advancing age. In worms, Dr. Guarente proposes that SIR2 increases longevity by maintaining chromatin organization and silencing unstable parts of the genome that might otherwise be mutated or inappropriately expressed. Taken together, these findings link genome silencing with metabolic rate and may provide a mechanistic connection between calorie restriction and life extension.

Regulation of the Oxidative Stress Response and Life Span by the Mammalian Shc Gene

Speaker

Pier Giuseppe Pelicci, M.D., Ph.D.

Chair, Department of Experimental Oncology

European Institute of Oncology

Several gene mutations in invertebrates have been shown to extend life and enhance resistance to stresses such as ultraviolet light or reactive oxygen species. But no stress-regulating genes had been identified in mammals until Dr. Pelicci and his coworkers showed that targeted mutations of the p66shc gene had these effects in mice. Dr. Pelicci chairs the Department of Experimental Oncology at the European Institute of Oncology, Milan, and is also an Associate Professor of Oncology at the University of Parma. He received his M.D. and Ph.D. from the University of Perugia and held postdoctoral fellowships at Istituto di Clinica Medica I, Institut National de la Sante et de la Recherche Medicale, and New York University Medical Center. In addition to receiving numerous awards, Dr. Pelicci serves on the editorial boards of journals published in Italy and the United States.

Topic

Regulation of the Oxidative Stress Response and Life Span by the Mammalian Shc Gene

One of the first clues about the regulation of mammalian stress response emerged from Dr. Pelicci’s laboratory, when he and his coworkers reported that targeted mutation of the mouse p66shc gene bolstered stress resistance and prolonged life. They view the p66shc protein, like other shc adaptor proteins, as one step along a signal transduction pathway that couples an activated tyrosine kinase receptor to Ras. The normal function of p66shc appears to be inducing apoptosis in cells undergoing oxidative stress.

When Dr. Pelicci’s team knocked out the p66shc gene in mice, fibroblasts from those animals exhibited increased resistance to oxidative and ultraviolet light damage. The animals themselves lived about one-third longer than normal mice, even though their calorie intake was not restricted. The researchers hypothesize that p66shc, which is concentrated in mitochondria, interferes with the metabolism of reactive oxygen species, allowing them to accumulate and trigger apoptosis. Thus, when the gene has been deleted, apoptosis is delayed or blocked.

The researchers wondered whether the price for living longer would be a greater susceptibility to tumors. But when they compared mice that were hetero- or homozygous for the p66shc deletion with control animals, they saw no increase in spontaneous or induced tumors in mice with the deletion. Future studies will home in on the exact role of p66shc in mitochondrial function.

Integrating the Cell-Death Pathway

Speaker

Stanley J. Korsmeyer, M.D.

Professor of Medicine

Dana-Farber Cancer Institute

In his pioneering studies of programmed cell death, Dr. Korsmeyer identified key genetic mechanisms that govern cell death and survival. These highly conserved genetic programs are crucial to embryonic development and later play important roles in the maintenance of the mature organism. His recent work goes beyond genetics to explore the biochemistry of cell death. Dr. Korsmeyer is the Director of the Program in Molecular Oncology at the Dana Farber Cancer Institute in Boston and Professor of Medicine at Harvard Medical School. He received a B.S. in biology and his M.D. from the University of Illinois. Dr. Korsmeyer is a member of the National Academy of Sciences and an Investigator at the Howard Hughes Medical Institute. His recent awards include the first American Society for Clinical Investigation Award and the Charles S. Mott Prize of the General Motors Cancer Research Foundation.

Topic

Integrating the Cell-Death Pathway

Deprived of external messages that tell them to keep dividing, or bombarded by signals that say they have outlived their usefulness, cells activate an intracellular suicide program. Early in this pathway, there is a checkpoint where the life-or-death decision is made. Much of Dr. Korsmeyer’s recent work focuses on the action at this checkpoint, where anti-apoptotic proteins such as BCL-2 square off against pro-apoptotic proteins including BAX, BAD, and BID.

When a death signal is received from the cell surface, the BAX protein, which has been drifting in the cytosol, pairs up with another of its kind, relocates to the mitochondrial membrane, and initiates several chains of events that lead to cell death, including caspase activation and mitochondrial dysfunction. If an anti-apoptotic protein like BCL-XL is administered in time, these events can be derailed. In a series of experiments in yeast, timely doses of BCL-XL were able to rescue some mitochondria that appeared to be dead, but were actually only stunned.

The more that Dr. Korsmeyer’s team learns about the tug of war between pro- and anti-apoptotic proteins, however, the more complex the story becomes. Recent experiments have been exploring the multiple roles of the BH3 domain of BID, which appears to pass along survival messages in some circumstances and death signals in others. Because these types of pathways appear to be involved in neurodegenerative diseases such as Alzheimer’s and Parkinson’s, Dr. Korsmeyer hopes that his investigations will ultimately lead to advances in diagnosis and treatment.

The Proteases to Die For

Speaker

Junying Yuan, Ph.D.

Associate Professor

Department of Cell Biology, Harvard Medical School

In 1993, Dr. Yuan broke new ground in apoptosis research by identifying both a cell-death gene in C. elegans and its mammalian homologues. This gene product, called Ced-3, was the first member of the caspase family of proteases – a group of enzymes now recognized as ubiquitous in programmed cell death. More recently, her laboratory in the Department of Cell Biology at Harvard Medical School has uncovered a direct link between a caspase and Alzheimer’s disease. Dr. Yuan has been an Associate Professor at HMS since 1996, having joined the faculty in 1992. Her investigations of cell death mechanisms in the nematode C. elegans began when she was a graduate student in the laboratory of Dr. H. Robert Horvitz at the Massachusetts Institute of Technology. She received her Ph.D. in Neuroscience from Harvard University in 1989.

Topic

The Proteases to Die For

While C. elegans has only one known caspase, humans have at least 14 of these cysteine proteases, now widely regarded as critical mediators of apoptosis signal transduction and execution. Caspases can be activated by apoptotic signals that target different cell compartments, such as death receptors located on cytoplasmic membrane, DNA damage to nuclei, free radical insult of mitochondria, and the build-up of proteins in the endoplasmic reticulum. Experiments in Dr. Yuan’s laboratory showed that the latter stimulus, known as ER stress, specifically activates caspase-12, an odd duck in the caspase family because it concentrates in the ER instead of floating in the cytosol with its relatives.

One well-known cause of ER stress in neurons is the accretion of amyloid-beta, the insoluble protein that forms plaques in the brains of patients with Alzheimer’s disease. As A-beta causes brain cells to die, a byproduct of their deaths is a caspase that cleaves amyloid precursor protein, thereby producing more of the deadly A-beta. Looking at this situation, and knowing that they had found an ER-specific caspase, Dr. Yuan and her colleagues wondered if there might be a promising connection between the two. When they exposed caspase-12 deficient cortical neurons to A-beta, the cells were resistant to A-beta toxicity. Dosing the neurons with antisense caspase-12 also reduced apoptosis. Experiments with caspase-12 knockout mice further supported a link between A-beta and this ER-specific caspase. Clearly a caspase-12 inhibitor would be a promising antidote to the neurodegeneration that occurs in Alzheimer’s disease, Dr. Yuan said.

The Genetic Basis for Neurodegeneration in Alzheimer’s Disease

Speaker

Rudolph E. Tanzi, Ph.D.

Professor of Neurology and Neuroscience

Department of Neurology, Massachusetts General Hospital

Alzheimer’s disease (AD) is the most common cause of dementia in elderly people, and Dr. Tanzi has been instrumental in discovering genes that account for nearly half of early-onset familial Alzheimer’s disease (FAD). In addition to analyzing the harm these genes do in early-onset FAD, he and his colleagues are also exploring the complex genetic underpinnings of the more common, late-onset form of Alzheimer’s. Dr. Tanzi is Director of the Genetics and Aging Unit in the Department of Neurology at Massachusetts General Hospital, as well as Professor of Neurology and Neuroscience at Harvard Medical School. He received his B.S. in microbiology and history from the University of Rochester and his Ph.D. in Neurobiology from Harvard University. Dr. Tanzi has received numerous honors, including the Metropolitan Life Award for Medical Research and the Potamkin Prize.

Topic

The Genetic Basis for Neurodegeneration in Alzheimer’s Disease

Dr. Tanzi made history in 1987 with the discovery of the amyloid-beta precursor gene, the first genetic cause of familial Alzheimer’s disease. He later played an important role in the discovery of two other important genes for early-onset FAD, called Presenilin 1 and 2, and since then his lab has been investigating how these gene defects cause neurodegeneration in AD, including their roles in apoptosis and oxidative stress. In addition, Dr. Tanzi has been seeking genetic risk factors for the more common, late-onset form of AD. Toward this end, he and colleagues at the National Institutes of Health carried out a high-resolution genome screen of more than 400 families.

Prior to this screen, the only known risk factor for late-onset AD was APOE4, one of three alleles for a gene that directs production of apolipoprotein E. People with two copies of the APOE4 polymorphism are at especially high risk for developing AD; people with one copy are at somewhat elevated risk. The genome-wide screen turned up another, unrelated polymorphism that may pack the same wallop as a double dose of APOE4. This is A2M-2, an allele of the gene for alpha2 macroglobulin (A2M). The A2M-2 variant is carried by about 30% of the U.S. population, and Dr. Tanzi and his colleagues estimate that people with this allele are three times more likely to develop late-onset AD than those with standard A2M.

The researchers are now trying to discover whether the FAD mutations and the polymorphisms linked to late-onset AD might, in the final analysis, use the same basic strategies to cause disease. One promising hypothesis is that they may alter the cell’s ability to handle calcium, and that this in turn may promote the formation of damaging amyloid-beta plaques. If confirmed, this could pave the way to new therapeutic approaches.