During embryonic brain development, reelin is secreted by Cajal-Retzius (CR) cells into the extracellular matrix (ECM) and serves to orchestrate the positioning of migrating neurons in the developing cortex, hippocampus and cerebellum. In the cortex of adult rat, reelin is secreted from a subpopulation of GABAergic neurons into the ECM by a constitutive mechanism. In adult non-human primates, reelin, present in the ECM, is contiguous with alpha3-containing integrin receptors in the vicinity of dendritic spines. This raises the possibility that adult reelin-integrin interactions may contribute to synaptic plasticity by modulating the expression of cytoskeletal proteins that facilitate the trophism of dendritic spines. In the post-mortem brains of schizophrenia patients, basal dendritic spine density is decreased. Moreover, in brains obtained from patients diagnosed with schizophrenia and bipolar illness with psychosis, reelin mRNA and protein levels are reduced by approximately 50 percent. Cytosine methylation of genomic DNA influences a panorama of cellular processes, including gene transcription, genomic imprinting, and genome stability. Experiments outlined in this application will expand upon our preliminary data that support our hypothesis that the human reelin promoter is modulated by DNA methylation, which determines transcription factor accessibility through alterations in chromatin structure. Our goal is to obtain information relevant to mechanisms responsible for appropriate temporal and spatial patterns of reelin gene expression. We will define those parts of the promoter operative in modulating reelin expression in primary neuronal cell cultures (cortical neurons and glia and cerebellar granule neurons). Using this information, we will identify trans-acting factors that interact with these regulatory regions (Aim 1). Secondly, we will also focus on the role that DNA methylation plays in defining reelin gene (RELN) expression in neuronal precursors differentiating in vitro (Aim 2). Finally, we will generate transgenic animals using the reelin promoter to drive expression of a lacZ reporter gene and examine the role of these regulatory sequences in targeting neuronal expression of RELN (Aim 1). We will examine the methylation pattern of the human promoter transgene in various primary cultures derived from these transgenic mice (Aim 2) and will compare the patterns of methylation of the endogenous gene with the human reelin transgene. Information obtained from the proposed experiments will provide the framework for formulating hypotheses relevant to mechanisms by which reelin expression may be functionally compromised in psychiatric diseases. Ultimately, an understanding of events responsible for modulating reelin expression may provide preliminary clues as to how the gene might be manipulated in the future as one potential therapeutic approach to these complex mental disorders.
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