Genetic and alcohol regulation of brain RNA levels

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DESCRIPTION (provided by applicant): The goal of this proposal is to provide Dr. Susan Bergeson career development opportunities including mentoring by Dr. Adron Harris, microarray training with Dr. Vishy Iayer, and a reduction in teaching effort that will allow time to develop an independent research program. The excellent research environment, faculty and facilities of the University of Texas will allow expansion of her expertise in molecular biology toward a new and innovative analysis of the role of gene expression in alcohol tolerance and dependence. The aims of the research component of the proposal are to 1) Identify changes in brain MRNA levels produced by acute alcohol exposure and to 2) Utilize mouse behavioral genetics to link I coordinated changes in expression to adaptations in molecular neurocircuitry that cause tolerance, dependence and sensitization. This will be accomplished by MRNA differential display (DD) coupled with DNA microarray analysis. Genes within known Quantitative Trait Loci (QTL) will be given priority and analysis of congenic mice constructed for acute withdrawal QTLs will provide a proof-of-concept. QTL analysis is currently a common method for mapping chromosomal regions that contain genes important in complex (polygenic) mammalian traits, including responses to alcohol but is only a first step in understanding mechanisms underlying alcohol's effects. Several expression studies, nave identified genes whose mRNA levels are changed by alcohol's actions yet no clear picture of the molecular, consequences of alcohol action have emerged. Combining genetic tools and molecular techniques offers a logical strategy for the detection and validation of important genetic differences that influence alcohol-related traits. A modified DD analysis, which has the capacity to screen most expressed transcripts as well as identify insertion and deletion polymorphisms, will be completed on C57Bl/6J (B6) and DBA/2J (D2) mice and two congenic strains created as a result of QTL analysis for acute alcohol withdrawal. First, DD analysis of B6, D2, and the congenic strains will provide alcohol-responsive and strain specific expression differences. Next, microarrays will be used for validation, and dose-effect, time-course, and brain region specificity will be defined by adding the DD isolated genes as additional targets for DNA microarrays analysis providing sufficient power to elucidate, by cluster analyses, molecular pathways with convergent regulation by ethanol. In addition, expression and sequence polymorphisms between B6, D2 and these congenic strains will be of value to numerous studies that have or will use a BXD strategy. All genetic and profiling data will be freely shared by deposition in the MGI data base (http:/www.informaticsjax.org) and a web site created at UT that details both the protocol and results; for an example see: V.Iyer,(http://genome-www.stanford.edu/serum).