Michael P.

Lab research focus: My lab has recently identified a calcium regulator which is translated at neuronal synapses. We have characterized its subcellular localization and identified pharmacological stimuli that can regulate its synthesis. In addition to elucidating the molecular mechanism by which it regulates calcium, we have also linked its translation to specific mouse models of cognitive disorders. This work, (Oyang et al., 2011) was recently published in PLoS One (http://dx.crossref.org/10.1371%2Fjournal.pone.0024879). Methods used: I supervised a 6 member lab utilizing an array of model systems including in vivo rodent models, primary rodent neuronal cultures, tumor cell lines, and in vitro assays. Techniques we employ include immunocytochemistry, immunohistochemistry, fluorescent in situ hybridization, pharmacological manipulations, live cell calcium imaging, confocal microscopy, quantitative PCR, and protein and nucleic acid biochemistry. We also maintained an independent rodent breeding colony. Responsibilities: As a supervisor, I was responsible for: Directing the overall research focus of the lab, forming collaborations with faculty within and outside the college, overseeing the lab budget, training students and supervising research projects

•Project summary: I discovered a novel molecular mechanism for retinoic acid (RA) mediated translational control at hippocampal synapses via the retinoic acid receptor alpha (RARa, and played a key role in identifying a new function for RA during adult synaptic plasticity. This work was published in PNAS (Poon and Chen, 2008; Maghsoodi et al., 2008) and Neuron (Aoto et al., 2008) • Novel methods developed: Starting with protein and RNA prediction software, I identified a putative RARa RNA binding domain and tested it experimentally by devising a novel hybrid in vitro/in vivo interaction screen using recombinant protein domains and brain-derived mRNA targets Designed a transfectable, cell-based sensor to monitor levels of retinoic acid (RA), a photosensitive and labile molecule, in living neurons • Advanced techniques: Applied a recombinant RARa-based selection of high affinity RNA species from a pool of random 30mer RNA sequences (SELEX) to identify RARa binding RNA sequence motifs Immunoprecipitated UV-crosslinked RARa-RNA complexes from brain tissue followed by a SELEX-based candidate PCR approach to identify in vivo RARa binding mRNA targets •Other methods employed: Nitrocellulose filter binding assays for in vitro protein-RNA interaction; transfection-based assay for assessing translational control in live cells; use of pharmacological agonists and antagonists to examine learning-related changes in cell culture and synaptoneurosomes; bred and maintained a transgenic mouse line for cellular and molecular studies

•Project summary: Using a method for isolating neuronal dendrites that I pioneered and rigorously tested, I identified 19 novel mRNAs by in situ hybridization, more than doubling the number of species known at the time. This method enabled us to study the molecular mechanisms of RNA localization and translation at the synapse. This work was published in J. Neurosci (Poon et al., 2006). •Novel methods created: Harvesting dendritic mRNA by plating cultured hippocampal neurons onto filters containing 3micron pores through which dendrites, but not cell bodies, could pass through and be isolated; method was validated by gene expression profiling •Key accomplishments: Generated microarray data and cDNA libraries from nanogram amounts of dendritic mRNA; confirmation by quantitative PCR and fluorescent in situ hybridization Detection of rare dendritic protein/mRNA complexes by confocal microscopy using combined fluorescent in situ hybridization and immunocytochemistry •Other methods employed: Used pharmacological reagents to induce or inhibit synaptic plasticity in cultured neurons to query synaptic mRNA localization and translation; used statistical analysis of microarray data to identify subpopulations of dendritic mRNAs; developed lab protocol for generating primary rat hippocampal cell cultures, designed and validated an antibody for the protein neuronatin

Advisor: Kelsey Martin; Dissertation titled: Identification of Localized mRNAs in Dendrites of Cultured Hippocampal neurons

1995-1998 Undergraduate researcher, Gabriele Ronnett lab, Johns Hopkins University School of Medicine