HomeResearchMolecular Mechanisms Regulating Neuronal Phenotype

Our Scientists

Molecular Mechanisms Regulating Neuronal Phenotype

Research Summary

Gail Mandel studies the molecular mechanisms underlying regulation of gene expression and function in the nervous system. A major effort is in understanding how transcriptional regulators, particularly repressors, guide transitions between developmental stages. Another focus of the lab is aimed at elucidating the basis of neuronal:glial communication and how this communication is altered in neurological diseases.

A central goal in neuroscience is to determine the genetic basis of neurological disorders. Many of these pathological states result from defects in gene regulatory programs that are fundamental to all cell types but lead to dysfunction specifically within the nervous system. My lab is investigating the basis of this phenomenon and has identified regulatory programs involving cell-cell interactions between neurons and glia as causal to certain pathological states of brain development. We have corrected, in a mouse model, the pathological, physiological, and behavioral states of one neurological disorder, Rett syndrome (RTT), by fixing the defective gene in the astrocytes. We continue to explore the mechanisms underlying normal neuronal-specific gene regulation as well as the cell-cell interactions crucial for neuronal signaling that are aberrant in disease.

Figure 1: Transcription factor REST...

Populations of Cells Expressing an Embryonic Gene Within the Adult Brain
Our previous studies showed that the genetic program responsible for determining a neuronal cell fate early in development is distinct from the program that activates genes required for later neuronal functions such as synaptogenesis. Specifically, we identified an important transcription factor, which we termed REST, that regulates the expression of genes required for terminal differentiation. REST coordinately represses, in embryonic stem cells and neural progenitors, thousands of genes required for metabolic function and electrical excitability in mature neurons. The loss of REST, timed to occur just at terminal differentiation, triggers expression of the suite of genes that define the mature neuronal phenotype. No other transcription factor is better associated with acquisition of the mature neuronal phenotype. Indeed, recent work from other investigators showed that REST is a central player in human cortical development and is implicated in diseases of the cortex, such as microcephaly. It came as a surprise, therefore, when we discovered, by developing better antibodies for examining REST expression in vivo, that some neurons in the mature brain reexpress REST later in life. These neurons are in discrete brain regions that are involved in specialized functions. For example, in the cerebellum, we identified a small fraction of neurons that express REST at high levels. We are developing methods to determine why this limited population of cells expresses a gene whose function is associated primarily with stem/progenitor cells. Outside the nervous system, REST functions as a tumor suppressor. Further studies of these two disparate REST functions, outside and inside the nervous system, may provide insights into roles for neuronal genes in tumor biology.

Rett Syndrome: A Complex Neurological Disorder Involving Interactions Between Neurons and Astrocytes
RTT is a neurological disorder that affects 1 in 10,000–15,000 births of girls. Afflicted individuals exhibit outwardly normal development for 12–18 months; then they begin to regress, losing verbal communication and purposeful hand motions. They have smaller brains than normal, they acquire severe respiratory problems, and they often have seizures. The disease is due to sporadic mutations in the methyl DNA binding protein MeCP2, which is located on the X chromosome. As a result, and because of dosage compensation in mammals, girls are mosaics of approximately half normal and half mutant cells throughout their body. Mouse models for RTT have revealed that loss of MeCP2 from neurons contributes significantly to the disease in mice, and there are similarities in the phenotypes of afflicted mice and humans. Despite these findings, as yet no single gene expression change as a result of the absence of MeCP2 has been shown to account for RTT. We found recently that loss of MeCP2 from glia, a non-neuronal supporting cell type, is a contributory factor to RTT and, surprisingly, fixing glia alone can dramatically improve survival and behavior in mouse models. Our work now focuses on determining the precise mechanisms underlying normal and abnormal interactions between specific types of glia and neurons. For example, as part of our long-term collaboration with physiologist Paul Brehm, using dual patch recordings we have identified chemical signaling between astrocytes and neurons. The nature of the chemical communication, and whether these modes of communication are affected in RTT cells, is an active area of investigation in my lab.

Movie 1: Array tomographic image of nucleus of neuron in a female mosaic mouse deficient in MeCP2. Green, anti-GFP in MeCP2-GFP knock-in mouse; red, anti-histone H3K9me3; blue, DAPI stain for heterochromatin.

Knock-in mouse: Jackson Laboratory; staining and image: Michael Linhoff.

We are also using high-resolution array tomography to test the hypothesis that loss of MeCP2 causes global changes in chromatin conformation. Such global changes could have the effect of altering thousands of genes so that the homeostasis of affected cells is disrupted. This type of scenario would be consistent with the observation that many mRNAs are both up- and downregulated upon loss of MeCP2. The movie shows a nucleus in a mutant mosaic female RTT mouse brain. The nucleus is in a wild-type cell because of inactivation of the X chromosome that contains the mutated MeCP2 allele. The nucleus is stained for MeCP2 (green) and a histone modification, H3K9me3 (red), both of which are known to be associated with heterochromatin (4',6-diamidino-2-phenylindole, blue). We have detected differences in chromatin between wild-type and mutant cells in mosaic brains from female RTT mice and are characterizing these differences. We are also combining array tomography with fluorescent in situ hybridization to look at the relationship of chromatin modifications and specific DNA sequences with respect to the MeCP2 location.

Exploring Ways to Fix RTT Mutations
In collaboration with a team of scientists at Vollum and the University of Puerto Rico, we are pioneering an approach to fix disease-causing mutations by using mice modeling a class of human RTT mutations as a test case. The approach harnesses the natural ability of site-specific RNA editing enzymes that can change adenosines to guanosines in RNA. When used in conjunction with a small antisense guide RNA, we hope to specifically target adenosines that were mutated from guanosines in the Mecp2 gene. Because RNA editing does not change other properties in the mRNA, this approach has the advantage that the correctly edited mRNA, and thus the encoded MeCP2 protein, remains at a normal physiological level in the cell. We are testing this approach to rescue mutations in mammalian and zebrafish systems and using combinations of behavior, immunohistochemistry, live imaging, and physiology to determine the extent of rescue and distribution of rescued cells within the nervous system.

Grants from the National Institutes of Health and Rett Syndrome Research Trust provided partial support for these projects.

As of February 27, 2014

Scientist Profile

Investigator
Oregon Health & Science University
Molecular Biology, Neuroscience