Abstract

The broad, long-term objectives of this project are to gain novel knowledge about a highly specialized calcium (Ca2+) channel - the inositol (1,4,5)-trisphosphate receptor (InsP3R). The proposed studies are fundamental to understanding Ca2+ signaling and may provide information relevant to multiple human diseases. My laboratory established functional expression of rat InsP3R1 in Sf9 cells and characterized properties of wild type and several mutant forms of recombinant InsP3Rl. We will continue to take advantage of the unique opportunity to measure functional activity of recombinant InsP3R and extend our structure-function analysis to other InsP3R isoforms.
The specific aims of the proposal are: 1. To compare functional properties of different InsP3R isoforms in identical experimental conditions. The rat InsP3R1, InsP3R2, InsP3R3, D. melanogaster, and C. elegans InsP3R isoforms will be expressed in Sf9 cells and characterized in planar lipid bilayers. 2. To identify structural determinants responsible for the InsP3R1 modulation by Ca2+. Ca2+-binding and functional experiments will be performed to search for novel InsP3R1 Ca2+ sensor mutants. Ca2+ modulation of InsP3R-1/3 and lnsP3R-3/1 chimeric proteins will be analyzed in planar lipid bilayers to identify the locations of Ca2+-modulatory sites in the InsP3R sequence. 3. To characterize InsP3R modulation by PKA phosphorylation. Effects of PKA on InsP3R1 SII(+/-) isoforms, InsP3R2, and InsP3R3 channels will be evaluated. The PKA phosphorylation sites in InsP3Rsequence will be mutated and the sensitivity of generated mutants to PKA modulation will be analyzed. InsP3R1-PKA association will be investigated in biochemical experiments. 4. To determine an importance of InsP3R1 modulation by PKA in vivo. Dopamine-induced responses in striatal medium spiny neurons will be studied by Ca2+ imaging. Experiments with striatal neurons from DARPP-32 and D2 receptor knockout mice will be performed.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS038082-07
Application #
7010650
Study Section
Neurotransporters, Receptors, and Calcium Signaling Study Section (NTRC)
Program Officer
Stewart, Randall R
Project Start
1998-12-03
Project End
2009-01-31
Budget Start
2006-02-01
Budget End
2007-01-31
Support Year
7
Fiscal Year
2006
Total Cost
$347,214
Indirect Cost

  Institution

Name
University of Texas Sw Medical Center Dallas
Department
Physiology
Type
Schools of Medicine
DUNS #
800771545
City
Dallas
State
TX
Country
United States
Zip Code
75390

  Publications

Bezprozvanny, Ilya (2013) Reconstitution of endoplasmic reticulum InsP3 receptors into black lipid membranes. Cold Spring Harb Protoc 2013:
Bezprozvanny, Ilya (2013) Bilayer measurement of endoplasmic reticulum Ca2+ channels. Cold Spring Harb Protoc 2013:
Bezprozvanny, Ilya (2013) Preparation of microsomes to study Ca2+ channels. Cold Spring Harb Protoc 2013:
Kasumu, Adebimpe W; Liang, Xia; Egorova, Polina et al. (2012) Chronic suppression of inositol 1,4,5-triphosphate receptor-mediated calcium signaling in cerebellar purkinje cells alleviates pathological phenotype in spinocerebellar ataxia 2 mice. J Neurosci 32:12786-96
Kasumu, Adebimpe; Bezprozvanny, Ilya (2012) Deranged calcium signaling in Purkinje cells and pathogenesis in spinocerebellar ataxia 2 (SCA2) and other ataxias. Cerebellum 11:630-9
Kasumu, Adebimpe W; Hougaard, Charlotte; Rode, Frederik et al. (2012) Selective positive modulator of calcium-activated potassium channels exerts beneficial effects in a mouse model of spinocerebellar ataxia type 2. Chem Biol 19:1340-53
Bezprozvanny, Ilya (2011) Role of inositol 1,4,5-trisphosphate receptors in pathogenesis of Huntington's disease and spinocerebellar ataxias. Neurochem Res 36:1186-97
Bezprozvanny, Ilya (2010) Inositol 1,4,5-tripshosphate receptor, calcium signaling, and polyglutamine expansion disorders. Curr Top Membr 66:323-41
Bezprozvanny, Ilya; Klockgether, Thomas (2009) Therapeutic prospects for spinocerebellar ataxia type 2 and 3. Drugs Future 34:
Liu, Jing; Tang, Tie-Shan; Tu, Huiping et al. (2009) Deranged calcium signaling and neurodegeneration in spinocerebellar ataxia type 2. J Neurosci 29:9148-62

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