The broad goal of this project is to identify the molecular mechanisms that underlie ion permeation and channel gating and its regulation in inositol trisphosphate receptors (InsP3R). The InsP3R is a ubiquitous family of intracellular Ca2+ release channels that participates in the generation of complex Ca2+ signals that regulate many physiological processes, and that has been implicated in several human diseases. Nevertheless, the molecular mechanisms in the channel that regulate its ion permeation properties, gating (opening and closing) and regulation are poorly understood. Its intracellular location has generally limited studies of its single channel properties, and its ubiquitous expression has impeded the development of robust systems for recombinant mammalian isoform expression. A challenge is to understand the function and regulation of this family of ion channels in order to understand its roles in Ca2+ signaling in normal and disease states. We previously developed patch clamp electrophysiology of isolated nuclei as a powerful approach for studying the single channel properties of the InsP3R in its native endoplasmic reticulum (ER) membrane. During previous funding cycles, we have made considerable progress in understanding the molecular physiology of recombinant mammalian InsP3Rs. We have developed a robust system that now greatly facilitates electrophysiological characterization of recombinant InsP3R channels in a completely null background. Importantly, we developed novel molecular models that can account for diverse regulation of channel gating that we have previously identified. These models provide an invaluable framework for interpreting the molecular consequences of mutations introduced into the channel sequences. Our development of nuclear patch electrophysiology and novel cell systems for mammalian InsP3R expression now enables detailed studies of wild type and mutant channels in native ER membranes. By comparison with nearly all other ion channels, and despite its role in generating diverse Ca2+ signals in nearly all cells that are responsible for regulating numerous cell physiological processes, the molecular physiology of the InsP3R is fundamentally understudied. Furthermore, the InsP3R has been linked to human disease processes, including apoptosis, neurodegeneration, cerebellar ataxias, and cardiovascular disease, and it may represent a therapeutic target in others. Thus, there are compelling reasons to understand the detailed properties of this ion channel. This knowledge could also be important to the development of therapeutic agents that may target InsP3R. Thus, this project could have a major impact. We propose three specific aims to characterize the molecular mechanisms that contribute to permeation, gating and channel regulation. The results of these studies should provide new insights into the molecular physiology of the family of InsP3R Ca2+ release channels.

Public Health Relevance

Inositol trisphosphate receptors are a family intracellular calcium release channels that generate diverse calcium signals that regulate numerous cellular processes and are altered in various human diseases. Nevertheless, nearly nothing is known about the molecular bases within these large complex proteins that mediate their ion channel functions. The studies proposed here will discover the molecular bases for ion permeation and regulation of channel opening and closing. This knowledge is critical for understanding inositol trisphosphate receptor function and it is important clinically, because receptor function is disrupted in diseases such as Alzheimer's disease, spinocerebellar ataxia and cardiovascular disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Mental Health (NIMH)
Type
Research Project (R01)
Project #
5R01MH059937-13
Application #
8225219
Study Section
Neurotransporters, Receptors, and Calcium Signaling Study Section (NTRC)
Program Officer
Asanuma, Chiiko
Project Start
1999-12-01
Project End
2015-02-28
Budget Start
2012-03-01
Budget End
2013-02-28
Support Year
13
Fiscal Year
2012
Total Cost
$396,000
Indirect Cost
$148,500
Name
University of Pennsylvania
Department
Physiology
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Joseph, J Donald; Peng, Yi; Mak, Don-On Daniel et al. (2014) General anesthetic isoflurane modulates inositol 1,4,5-trisphosphate receptor calcium channel opening. Anesthesiology 121:528-37
Shilling, Dustin; Müller, Marioly; Takano, Hajime et al. (2014) Suppression of InsP3 receptor-mediated Ca2+ signaling alleviates mutant presenilin-linked familial Alzheimer's disease pathogenesis. J Neurosci 34:6910-23
Siebert, Adam P; Ma, Zhongming; Grevet, Jeremy D et al. (2013) Structural and functional similarities of calcium homeostasis modulator 1 (CALHM1) ion channel with connexins, pannexins, and innexins. J Biol Chem 288:6140-53
Taruno, Akiyuki; Vingtdeux, Valerie; Ohmoto, Makoto et al. (2013) CALHM1 ion channel mediates purinergic neurotransmission of sweet, bitter and umami tastes. Nature 495:223-6
Shilling, Dustin; Mak, Don-On Daniel; Kang, David E et al. (2012) Lack of evidence for presenilins as endoplasmic reticulum Ca2+ leak channels. J Biol Chem 287:10933-44
Kopil, Catherine M; Vais, Horia; Cheung, King-Ho et al. (2011) Calpain-cleaved type 1 inositol 1,4,5-trisphosphate receptor (InsP(3)R1) has InsP(3)-independent gating and disrupts intracellular Ca(2+) homeostasis. J Biol Chem 286:35998-6010
Muller, Marioly; Cheung, King-Ho; Foskett, J Kevin (2011) Enhanced ROS generation mediated by Alzheimer's disease presenilin regulation of InsP3R Ca2+ signaling. Antioxid Redox Signal 14:1225-35
Muller, Marioly; Cardenas, Cesar; Mei, Lijuan et al. (2011) Constitutive cAMP response element binding protein (CREB) activation by Alzheimer's disease presenilin-driven inositol trisphosphate receptor (InsP3R) Ca2+ signaling. Proc Natl Acad Sci U S A 108:13293-8
Vais, Horia; Foskett, J Kevin; Mak, Don-On Daniel (2011) InsP3R channel gating altered by clustering? Nature 478:E1-2; discussion E2-3
Foskett, J Kevin (2010) Inositol trisphosphate receptor Ca2+ release channels in neurological diseases. Pflugers Arch 460:481-94

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