Increases in cytoplasmic Ca2+ evoked by inositol(1,4,5)-trisphosphate receptors (IP3Rs) regulate many physiological events. Dysfunction of the IP3-stimulated Ca2+ release pathway is involved in neurodegenerative and neurological disease, as well as exocrine, autoimmune and kidney disorders, cardiac abnormalities and cancer. To appreciate how cellular functions are controlled by Ca2+ signals, and how pathological aberrations subvert the IP3R signaling pathway, we must understand how the distribution and properties (the `functional architecture') of IP3Rs control the spatiotemporal organization of cellular Ca2+ signals. Here, we will resolve the properties of IP3Rs at three distinct levels of organization from single molecules in vitro (Aim 1) to local, subcellular (Aim 2) and global, tissue-level IP3R architecture in live embryos (Aim 3). We will address: (1) How, for individual IP3R, does a single molecule of IP3 bind to the IP3R? (2) How is IP3R function modulated within the dynamic context of endoplasmic reticulum structures where they reside? (3) How does a changing global complement of Ca2+ channels and pumps at a regional level in a developing vertebrate embryo impacts the patterning of Ca2+ signals and cell function. To address these challenges, we have optimized: (i) a novel single molecule imaging approach competent to resolve the properties of individual IP3Rs (Aim 1), (ii) a dual confocal microscope to simultaneously resolve ER architecture and the functionality of IP3Rs (Aim 2) and (iii) designed tools and reagents to probe the distribution and role of key families of Ca2+ channels and pumps during vertebrate embryogenesis (Aim 3). The broad significance of this work is in understanding principles controlling ion channel dynamics and thereby the spatial kinetics of Ca2+ signals that control unitary, cellular and systems-level responses. Such data will aid our understanding of the role of ubiquitous Ca2+ signaling pathways in health and disease.

Public Health Relevance

The goal of this project is to understand how particular cell signaling events are organized within cells by understanding first how proteins work at the unitary level in vitro and then appreciating how these proteins work when organized within cells and within developing embryos in vivo. The goal is to provide deeper understand of how intracellular Ca2+ channels behave and how pathological events subvert their function.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM088790-08
Application #
8214675
Study Section
Neurotransporters, Receptors, and Calcium Signaling Study Section (NTRC)
Program Officer
Gindhart, Joseph G
Project Start
2004-07-01
Project End
2014-01-31
Budget Start
2012-02-01
Budget End
2014-01-31
Support Year
8
Fiscal Year
2012
Total Cost
$304,313
Indirect Cost
$100,942
Name
University of Minnesota Twin Cities
Department
Pharmacology
Type
Schools of Medicine
DUNS #
555917996
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
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Lin-Moshier, Yaping; Marchant, Jonathan S (2013) The Xenopus oocyte: a single-cell model for studying Ca2+ signaling. Cold Spring Harb Protoc 2013:
Subramanian, Veedamali S; Nabokina, Svetlana M; Patton, Joseph R et al. (2013) Glyoxalate reductase/hydroxypyruvate reductase interacts with the sodium-dependent vitamin C transporter-1 to regulate cellular vitamin C homeostasis. Am J Physiol Gastrointest Liver Physiol 304:G1079-86
Lin-Moshier, Yaping; Marchant, Jonathan S (2013) Nuclear microinjection to assess how heterologously expressed proteins impact Ca2+ signals in Xenopus oocytes. Cold Spring Harb Protoc 2013:

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