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Session 3: Epigenetics in CNS Plasticity


Deborah Zelinsky
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Deborah Zelinsky

PART 1: CELLULAR AND MOLECULAR MECHANISMS OF MOTOR MEMORY By: Yue Yang, Ph.D., Department of Neurobiology at Northwestern University in Chicago

This discussion centered on the question of what molecular mechanism drives motor learning. It was determined that gene expression is important for neuron regulation in both early development and mature neurons, demonstrating the plasticity of both structure and function of chromatin. There is a specific order in proliferating vs. migrating vs. mature neurons. Calcium influx initiates a signaling cascade, triggering to start more genes to express, creating proteins that help restructure. Promoters and enhancers have a changing relationship during development in precursor cells as opposed to mature cells, affecting connectivity and changing genome regulation. Upon neurostimulation, gene enhancers are brought closer to gene promoters.

Cells studied were the anterior dorsal cerebellar vermis (ADCV) – where granular neurons have hundreds of genes that are expressed during a conditioned motor stimulus. Some genes are downregulated and others are upregulated, but the end result is that the chromatin remodels and is followed by changes in the genome architecture. Genome architecture is dynamic, leading to plasticity which rewires the neurological circuits. Experiments used a method of having antibodies to isolate gene promoters and label them, and see which part of a gene locus the promotor interacts with – usually the enhancer. Activation of the conditioned stimulus reorganizes gene enhancer and promotor interaction, thus showing the dynamic genome architecture.

Rather than using the antibodies, optogenetic stimulation demonstrated that the distance between enhancer and promoter changed to get closer or farther. Experiments worked with a head-fixed mouse and measured the avoidance (startle) reaction when a touch stimulus was introduced. The stimulus was coupled with lighting changes to create an associated conditioned response. The adaptive behavior was called Delay-Tactile startle conditioning and took 80% of the mice an average of 5 days to learn that association, because at first, the LED light did not elicit any startle response. More assessments were done to determine exactly where In the brain the link was that associated the light with the startle response. Researchers were looking at gene promoters that affect transcription and how they interact with gene enhancers (up to 500,000 base pairs) or a whole chromosome (hundreds of millions of base pairs). The researchers isolated a brain region important for plasticity and adaptive behavior, the section to acquire new memories.

Gene expression is important for plasticity and acquiring memories. The conditioned stimulus has more gene expression. Genome architectural mechanisms and interactions between promoter and enhancer plays powerful roles in learning and memory. Impaired acquisition of conditioned learning can also be affected by the induction of activity-dependent transcription. Learning can be impaired by disrupting transcription with a knockout of a transcription factor (in this case, Rad21). Doing the same experiment on the mice with a knocked out transcription factor only 40% could learn the conditioned startle response in 5 days.

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CONCLUSION:

Experience-dependent chromatin structural reorganization in cerebellar neurons influences motor learning. When the nuclear signaling is altered in the granule cell, chromatin remodeling is activated, so that the genome architecture forms loops, allowing the promoter and enhancer to come in close proximity, affecting RNA expression and changing motor learning circuitry and thus, behavior.

Edited by Deborah Zelinsky
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Deborah Zelinsky

PART 2: CELLULAR AND MOLECULAR MECHANISMS OF MEMORY By: Johannes Graff assistant professor at the Brain-Mind Institute, School of Life Sciences, part of the Federal Institute of Technology Lausanne in Switzerland

This lecture discussed the genetic and epigenetic interactions in patients who have Alzheimer’s Disease -- the most prevalent neurodegenerative disease worldwide. 50 million people have cognitive decline due to Alzheimer’s induced shrinkage of brain tissue. Cellularly their brains exhibit build up of extracellular amyloid plaques build up and intracellularly, tau hyperphosphoralated neurofibril tangles exist.

Studying Alzheimer’s Disease (AD) in the population can be based on risk factors (Genome Wide Associated Studies) or the research can be based on looking at DNA loci in Epigenetic studies or, both methods combined. The combination increases the statistical power that is lacking in the epigenetic studies, and increases the insights into mechanisms, by having more locus specificity that is lacking in the genome-wide studies.

When researching quantitative trait loci (QTL) to see if there were any sections of DNA associated with a person having Alzheimer’s, Dr. Graff’s group found one! He discovered that PM20D1 is a trait locus for Alzheimer’s disease, with a molecular marker of rs708727 as the SNP that modulates its methylation, and thus, its gene expression. PM20D1 = Metalloproteinase is found in the hippocampus, its amount is increased when stressors mimicking alzheimer’s are around, such as oxidative stress or amyloid beta fibrils to simulate plaques.

Methylation worsened the RNA upregulation and expression and didn’t allow for neuroprotection. When looking at people without Alzheimer’s, fewer have methylated PM20D1 promoter regions. In people with Alzheimers, when the PM20D1 promoter region was NOT methylated, the disease progression was delayed. When the PM20D1 was methylated, the progression of the disease was faster.

There are AA carriers and GG genotypes. When exposed to Amyloid Beta or ROS, the rs708727 SNP is shown to form a chromatin reconfigured loop only in GG carriers. That loop allows for the enhancer and the promoter region of the PM20D1 to get closer to each other and trigger gene expression of more PM20D1. This PM20D1 then clusters around the amyloid plaques, resulting in increased cell survival and enhanced memory. The clustering of PM20D1 around amyloid plaques forms sort of a shield for neuroprotection. Experiments showed increased cell survival and decreased amyloid levels in the hippocampus and improved memory performance in novel objects recognition tasks. A lessened presence of PM20D1 increased the amyloid plaques and worsened memory performance, not having that neuroprotective clustering around the plaques.

PM20D1 methylation depends on rs708727. The risk of Alzheimer’s is higher with a AA carrier than a GG carrier, because the PM20D1 chromatin reconfiguring into a loop occurs with GG carriers. The loop is needed for the promoter and enhancer to interact, increasing the amount of PM20D1. It correlates with the DNA methylation region of the PM20D1 promoter also with a reduction in RNA expression. GG carriers have a high expression of PM20D1’s promotor region, not any of the other regions. A second experiment verified this, where CTCF binding also showed a high experession of PM20D1 only in GG carriers. Those two independent studies on neurodegenerative diseases, including AD had one looking at the hippocampus via a methylation assay, the other looking the frontal lobe. Various methods (incuding a Chromatin Configuration Capture) demonstrated the same finding, that rs708727 acts as an enhancer for the PM20D1 promoter.

CONCLUSIONS: 1) PM20D1 is a disease relevant locus (QTL) for Alzheimer’s Disease and the shift in configuration in GG genotypes allows for the rs708727 SNP to help upregulate the RNA which induces a neuroprotective effect. 2) Methylation promotes a more rapid brain deterioration in Alzheimer’s disease. Among people with AD, more have methylated PM20D1. With methylated PM20D1 there is no chromatin reconfigured loop, therefore no enhancer/promoter interaction, therefore no upregulation of PM20D1, and no neuroprotection for the amyloid plaques. 3) PM20D1 methylation is genetically determined. GG carriers are less likely to develop Alzheimer’s Disease, showing how the epigenetics loci studies link with the Genome wide studies.

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