Intracellular calcium is a critical signaling molecule in the brain for many functions including neuronal development and synaptic plasticity. A family of Ca2+/calmodulin-regulated protein kinases (CaMKs) are key regulators of several of these events. During the past several years we identified regulatory functions of CaM-KK and its downstream effector CaMKI in the morphology and motility of axonal growth cones and basal axonal outgrowth, activity-dependent dendritic arborization and early-phase long-term potentiation. In the current application we will focus on two critical aspects of neuronal development, axon specification and formation of dendritic spines, for which we have preliminary evidence for involvement of CaMKK and CaMKI. We will analyze the activation state of CaMKK and CaMKI isoforms in different subcellular compartments of cultured hippocampal neurons using several approaches including FRET analysis. From proteomic analysis, we have evidence for the existence of a CaMKK signalsome containing beta-PIX and GIT1, and FIX is an in vitro substrate for CaMKI. The role of this signaling complex, as well as several other CaMKI substrates (Numb and stathmin 2) known to regulate the actin and microtubule cytoskelton, in axon specification and spinogenesis will be explored. Phosphorylation of these proteins by CaMKI in neurons will be studied using phospho-specific antibodies. These signaling pathways from CaMKI to axonogensis and spinogenesis will be defined using multiple, independent approaches including dominant-interferring and constitutively-active constructs, pharmacological inhibitors, and RNA interference. The primary experimental system will be cultured hippocampal neurons, but results will be extended into cultured hippocampal slices where possible. Identifying the fundamental signaling pathwaysthat modulate these essential neuronal functions is foundational to understanding normal and pathological brain functions including axon regeneration and mental retardation. Structural abnormalities in dendritic arborization and spine formation, both in human patients and in mouse models, are common to numberous forms of mental retardation including Fragile X, fetal alcohol, Down's and Rett syndromes as well as several evironmental causes of retardation. Thus, these studies have strong implications for underlying causes of mental diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM041292-22
Application #
7739444
Study Section
Synapses, Cytoskeleton and Trafficking Study Section (SYN)
Program Officer
Gerratana, Barbara
Project Start
1991-03-01
Project End
2011-11-30
Budget Start
2009-12-01
Budget End
2011-11-30
Support Year
22
Fiscal Year
2010
Total Cost
$365,904
Indirect Cost
Name
Oregon Health and Science University
Department
Neurosciences
Type
Schools of Medicine
DUNS #
096997515
City
Portland
State
OR
Country
United States
Zip Code
97239
Wayman, Gary A; Tokumitsu, Hiroshi; Davare, Monika A et al. (2011) Analysis of CaM-kinase signaling in cells. Cell Calcium 50:1-8
Davare, Monika A; Saneyoshi, Takeo; Soderling, Thomas R (2011) Calmodulin-kinases regulate basal and estrogen stimulated medulloblastoma migration via Rac1. J Neurooncol 104:65-82
Saneyoshi, Takeo; Fortin, Dale A; Soderling, Thomas R (2010) Regulation of spine and synapse formation by activity-dependent intracellular signaling pathways. Curr Opin Neurobiol 20:108-15
Davare, Monika A; Fortin, Dale A; Saneyoshi, Takeo et al. (2009) Transient receptor potential canonical 5 channels activate Ca2+/calmodulin kinase Igamma to promote axon formation in hippocampal neurons. J Neurosci 29:9794-808
Yang, Dongren; Kim, Kyung Ho; Phimister, Andrew et al. (2009) Developmental exposure to polychlorinated biphenyls interferes with experience-dependent dendritic plasticity and ryanodine receptor expression in weanling rats. Environ Health Perspect 117:426-35
Pessah, Isaac N; Seegal, Richard F; Lein, Pamela J et al. (2008) Immunologic and neurodevelopmental susceptibilities of autism. Neurotoxicology 29:532-45
Wayman, Gary A; Lee, Yong-Seok; Tokumitsu, Hiroshi et al. (2008) Calmodulin-kinases: modulators of neuronal development and plasticity. Neuron 59:914-31
Saneyoshi, Takeo; Wayman, Gary; Fortin, Dale et al. (2008) Activity-dependent synaptogenesis: regulation by a CaM-kinase kinase/CaM-kinase I/betaPIX signaling complex. Neuron 57:94-107
Wayman, Gary A; Impey, Soren; Marks, Daniel et al. (2006) Activity-dependent dendritic arborization mediated by CaM-kinase I activation and enhanced CREB-dependent transcription of Wnt-2. Neuron 50:897-909
Schmitt, John M; Guire, Eric S; Saneyoshi, Takeo et al. (2005) Calmodulin-dependent kinase kinase/calmodulin kinase I activity gates extracellular-regulated kinase-dependent long-term potentiation. J Neurosci 25:1281-90

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