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The Journal of Neuroscience, December 20, 2006, 26(51):13390-13399; doi:10.1523/JNEUROSCI.3432-06.2006
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Development/Plasticity/Repair
Identification of Process-Localized mRNAs from Cultured Rodent Hippocampal Neurons
Michael M. Poon,1 Sang-Hyun Choi,2 Christina A. M. Jamieson,3,4 Daniel H. Geschwind,2,3,5 andKelsey C. Martin5,6 1Interdepartmental Program in Neuroscience, 2Program in Neurogenetics, Department of Neurology, 3Department of Human Genetics, 4Department of Urology, 5Semel Institute for Neuroscience and Human Behavior, Department of Psychiatry and Biobehavioral Sciences, and 6Department of Biological Chemistry, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, California 90095
Correspondence should be addressed to Kelsey C. Martin, Gonda 3506/Brain Research Institute, University of California, Los Angeles, 695 Charles Young Drive, South Los Angeles, CA 90095-1761. Email: kcmartin{at}mednet.ucla.edu
The regulated translation of localized mRNAs in neurons provides a mechanism for spatially restricting gene expression in a synapse-specific manner. To identify the population of mRNAs present in distal neuronal processes of rodent hippocampal neurons, we grew neurons on polycarbonate filters etched with 3 µm pores. Although the neuronal cell bodies remained on the top surface of the filters, dendrites, axons, and glial processes penetrated through the pores to grow along the bottom surface of the membrane where they could be mechanically separated from cell bodies. Quantitative PCR and immunochemical analyses of the process preparation revealed that it was remarkably free of somatic contamination. Microarray analysis of RNA isolated from the processes identified over 100 potentially localized mRNAs. In situ hybridization studies of 19 of these transcripts confirmed that all 19 were present in dendrites, validating the utility of this approach for identifying dendritically localized transcripts. Many of the identified mRNAs encoded components of the translational machinery and several were associated with the RNA-binding protein Staufen. These findings indicate that there is a rich repertoire of mRNAs whose translation can be locally regulated and support the emerging idea that local protein synthesis serves to boost the translational capacity of synapses.
Key words: synaptic plasticity; microarray; hippocampus; protein synthesis; mRNA localization; in situ hybridization; dendrite
Received April 7, 2006; revised Nov. 14, 2006; accepted Nov. 15, 2006.
Correspondence should be addressed to Kelsey C. Martin, Gonda 3506/Brain Research Institute, University of California, Los Angeles, 695 Charles Young Drive, South Los Angeles, CA 90095-1761. Email: kcmartin{at}mednet.ucla.edu
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