Scientists have turned to a yeast model to screen for genetic modifiers of amyloid beta (Abeta) toxicity and have identified several genes known to be associated with Alzheimer disease (AD) and a few surprises that could help explain how the toxic protein exerts its toxicity in the brain. The finding may ultimately lead to new pathways to target treatment, several experts told Neurology Today.
For now, however, the discovery in yeast may help answer a puzzling question that has stumped AD scientists: What is the mechanism by which Abeta causes neuronal damage? Despite decades of research on Abeta no one really knows how Abeta protein leads to cell death.
This study, published in the Dec. 2 Science, suggests a mechanism that scientists had yet to consider: the Abeta protein interferes with endocytosis. Proteins that sit on the cell surface have to be degraded and turned over. Abeta seems to impair endocytosis and this process provides an important clue how Abeta causes neurotoxicity.
Susan Lindquist, PhD, a member of the Whitehead Institute for Biomedical Research who is professor of biology at the Massachusetts Institute of Technology (MIT) and a Howard Hughes Medical Institute investigator, led the study. Her laboratory pioneered work with yeast models of protein misfolding and aggregation, specifically alpha-synuclein in Parkinson disease. Yeast is a eukaryote organism that shares a lot of cell biology with humans.
Several years ago they introduced alpha-synuclein into yeast and it caused cells to dysfunction. Specifically, it perturbed Golgi trafficking. Successful with this model, Dr. Lindquist's colleague, Sebastian Treusch, PhD, introduced Abeta into yeast cells to look at the effects of Abeta toxicity. Dr. Treusch is one of the authors of the Science paper.
The scientists directed the Abeta peptide to the cell's secretory pathway, to recapitulate its trafficking in human cells. As hoped, it was toxic. The yeast did not grow as it normally does, and formed toxic oligomeric structures implicated in AD pathology. They then used a genome-wide overexpression screen to test more than 5000 yeast genes for the ability to affect Abeta toxicity. They went gene by gene, turning the expression up or down to see what it would take to make things better or worse.
They ended up with 40 yeast genes that had a hand in influencing Abeta toxicity. Of these genes identified in yeast 12 had clear human homologs — several of which had connections to AD.
One of the hits is on a gene called PICALM, short for phosphatidylinositol binding clathrin assembly protein. The gene hadbeen associated with AD risk but it wasn't known if it had an influence on Abeta toxicity. PICALM has a role in endocytosis, as do BIN1 (box-dependent-interacting protein 1) and CD2A (CD2-associated protein), which also popped up on their yeast screen for Abeta toxicity modifiers.
It turns out that Abeta impaired endocytic trafficking, which means that the cells have a hard time degrading receptors and their proteins at the cell membrane. “There is only so much real estate on the cell surface,” said study co-author Rudolph Tanzi, PhD, director of the Genetics and Aging Research Unit at the Massachusetts General Institute for Neurodegenerative Disease & Joseph P. and Rose F. Kennedy Professor of Neurology at Harvard Medical School. The genes identified in yeast were sent to Dr. Tanzi to identify any homologs to human AD genes. “It is critically important to manage cell surface proteins,” he said.
When they repeated the studies in other models — glutamatergic neurons of C. elegans and primary rat cortical neurons — the team also found the Abeta impaired endocytic trafficking of a plasma membrane receptor.
DR. SEBASTIAN TREUSC...Image Tools
They also discovered that overexpressing the PICALM homolog in yeast rescued the endocytosis dysregulation and prevented Abeta toxicity, a finding that could lead to novel treatments directed at PICALM. The yeast model can also be used to screen for potential AD drugs.
“Clathrin-mediated endocytosis is a critical point of vulnerability to Abeta,” the scientists wrote in the paper. “Neurons are particularly vulnerable to perturbations in the homeostasis of endocytosis, because they must constantly recycle both neurotransmitters and their receptors…Our yeast model provides a tool for identifying genetic leads, investigating their mechanisms of action, and screening for genetic and small-molecule modifiers of this devastating and etiologically complex disease.”
Dr. Treusch added: “Yeast, C-elegans and cortical neurons are useful in testing how these genes and their protein products actually modify the toxicity of Abeta and how mutations in these genes might influence the Alzheimer's process.”
They also looked through data from two large epidemiological studies of aging, cognition and AD and matched two loci identified in the yeast screen. “Our yeast screen connects multiple human AD risk factors and suggested risk factors to Abeta toxicity,” said Dr. Treusch, who has moved from MIT to Princeton University, where he is a post-doctoral researcher. Dr. Lindquist and her colleagues in Boston are now doing small molecule screening and testing other genes found in their research.
The researchers “show that Abeta toxicity can be modeled in yeast, and that yeast modifiers of that toxicity can powerfully inform our understanding of genetic players in the human disease,” Leeanne McGurk, PhD, and Nancy M. Bonini, PhD, of the department of biology at the University of Pennsylvania wrote in an accompanying commentary in Science. “This combined approach can provide insight into the biological processes and the basis for new therapeutic intervention.” Dr. Bonini's laboratory uses fruit flies to model protein misfolding.
Lon S. Schneider, MD, professor of psychiatry, neurology and gerontology at the University of Southern California (USC) and director of the Alzheimer's Disease Research and Clinical Center, added that “this kind of modeling is significant for understanding potential pathophysiological processes and for choosing drug targets. The model yields potential new targets. In turn, a prospective therapeutic can be first tested in simple model organisms.”
“I hope this study stimulates more work in this area,” said Berislav V. Zlokovic, MD, PhD, professor and chair of the department of physiology and biophysics director at the Center for Neurodegeneration and Regeneration in the Zilkha Neurogenetic Institute at the USC Keck School of Medicine. “This is a challenging area that is not insurmountable. It makes sense to prevent Abeta toxicity by blocking the effect of endocytosis.”
But another expert offered more tempered enthusiasm. “Basic science modeling is important but it is a very long way to the development of a drug that people can take,” said Howard Fillet, MD, executive director and chief scientific officer for the Alzheimer's Drug Discovery Foundation in New York City. “We have known about the apolipoprotein E4 gene for a long time and know so much about its role and its metabolism and we are not even close to developing a drug that targets it.”
Dr. Fillet added: “In my mind, you can't model a disease as complicated as Alzheimer's in an in vitro system but you can model a pathway. While it is interesting, we have to temper our enthusiasm for this kind of work.”
Hear more about what this yeast model and other new therapeutic approaches are yielding for advances in Alzheimer disease in an interview with Rudolph Tanzi, MD, director of the Genetics and Aging Research Unit at the Massachusetts General Institute for Neurodegenerative Disease & Joseph P. and Rose F. Kennedy Professor of Neurology at Harvard Medical School: http://bit.ly/AqMXdf