The objectives of the proposed research are the development of new molecular engineering technologies for large-scale gene expression analysis from single neurons, and applications of these technologies to identify and characterize genes that are involved in long-term synaptic plasticity and growth. We will combine research expertise in Chemistry, Engineering and Biology to pursue the research and development of the following new molecular engineering approaches: (i) Massive Parallel DNA Sequencing Chip System for digital gene expression analysis from single cells and cell compartments;and (ii) Novel Molecular Probes for Real-time monitoring of multiple mRNA species in living neurons and defined cellular microdomains. Each of these technologies will be rigorously tested and validated using the simpler memory-forming network of Aplysia, a unique model organism for neurobiology. As a "proof-of concept", we will focus on using these approaches for the identification of gene-regulatory networks underlying the learning-induced synaptic growth. Specifically, we will characterize a molecular cascade of events induced by serotonin, leading to the formation of new synapses and a long-term enhancement of synaptic strength also known as cellular manifestations of learning and memory mechanisms. The long-term goal of this project is to implement these new technologies to explore two fundamental brain mechanisms: (1) the molecular basis of neuronal growth;(2) the molecular signals controlling synapse-specific neuronal plasticity. Using the sensory neurons of the neuronal networks in Aplysia as an experimental model, we will study the role of asymmetric mRNA distribution in integrative functions and phenotypes of eukaryotic cells. We will use a hierarchical design to achieve structural resolution of single-cell profiling in a descending fashion, where a parallel genomic and functional analysis will be performed according to the following scheme: single neuron->single axon->single synapse. The gene expression profiling will be validated using a set of complementary approaches, correlated with functional imaging of selected mRNAs at functionally characterized neurons and synaptic terminals during various stages of 5-HT induced synaptic growth. The combined approach based on Chemistry, Engineering, and Neuroscience will be used to understand how neurons and synapses operate in the context of learning and memory. The technologies developed and the biological discoveries made in the project will have a broad impact in deciphering the molecular mechanisms of neurological disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS060762-04
Application #
8014902
Study Section
Special Emphasis Panel (ZRG1-MDCN-K (90))
Program Officer
Mamounas, Laura
Project Start
2008-02-01
Project End
2012-01-31
Budget Start
2011-02-01
Budget End
2012-01-31
Support Year
4
Fiscal Year
2011
Total Cost
$557,039
Indirect Cost
Name
Columbia University (N.Y.)
Department
Engineering (All Types)
Type
Schools of Engineering
DUNS #
049179401
City
New York
State
NY
Country
United States
Zip Code
10027
Zhu, Jing; Qiu, Chunmei; Palla, Mirkó et al. (2014) A Microfluidic Device for Multiplex Single-Nucleotide Polymorphism Genotyping. RSC Adv 4:4269-4277
Palla, Mirkó; Guo, Wenjing; Shi, Shundi et al. (2014) DNA sequencing by synthesis using 3'-O-azidomethyl nucleotide reversible terminators and surface-enhanced Raman spectroscopic detection. RSC Adv 4:49342-49346
Raveendra, Bindu L; Siemer, Ansgar B; Puthanveettil, Sathyanarayanan V et al. (2013) Characterization of prion-like conformational changes of the neuronal isoform of Aplysia CPEB. Nat Struct Mol Biol 20:495-501
Puthanveettil, Sathyanarayanan V; Antonov, Igor; Kalachikov, Sergey et al. (2013) A strategy to capture and characterize the synaptic transcriptome. Proc Natl Acad Sci U S A 110:7464-9
Kassabov, Stefan R; Choi, Yun-Beom; Karl, Kevin A et al. (2013) A single Aplysia neurotrophin mediates synaptic facilitation via differentially processed isoforms. Cell Rep 3:1213-27
Moroz, L L (2012) Phylogenomics meets neuroscience: how many times might complex brains have evolved? Acta Biol Hung 63 Suppl 2:3-19
Qiu, Chunmei; Kumar, Shiv; Guo, Jia et al. (2012) Mitochondrial single nucleotide polymorphism genotyping by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using cleavable biotinylated dideoxynucleotides. Anal Biochem 427:202-10
Kohn, Andrea B; Citarella, Mathew R; Kocot, Kevin M et al. (2012) Rapid evolution of the compact and unusual mitochondrial genome in the ctenophore, Pleurobrachia bachei. Mol Phylogenet Evol 63:203-7
Guo, Jia; Ju, Jingyue; Turro, Nicholas J (2012) Fluorescent hybridization probes for nucleic acid detection. Anal Bioanal Chem 402:3115-25
Qiu, Chunmei; Kumar, Shiv; Guo, Jia et al. (2012) Design and synthesis of cleavable biotinylated dideoxynucleotides for DNA sequencing by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Anal Biochem 427:193-201

Showing the most recent 10 out of 19 publications