1993 — 2004 |
Pfaffinger, Paul |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Functional Domains of K+ Channel Subunit Proteins @ Baylor College of Medicine
The physiological properties of a neuron and other cells of the body are strongly affected by the variety and abundance of potassium channels expressed in the cell. A large number of K+ channel genes are being cloned that apparently code for single subunits of multisubunit K+ channel proteins, but how are these subunit proteins targeted to form the variety of K+ channel proteins found in a cell. A specific question these experiments will address is whether subunit primary structures guide K+ channel assembly in forming the electrical properties of neurons. The Preliminary Results describe experiments that identify a domain of a Shaker Subfamily K+ channel protein (T1 domain) that appears to be a self organizing tetramerization center; tetramerization is thought to be a critical step in the formation of K+ channels by K+ channel subunit proteins. This T1 domain does not appear to interact with a K+ channel clone from another gene subfamily. The Specific Aims of this grant are: 1) Further Characterization of the Shaker Subfamily T1 N-Terminal Tetramerization Domain, 2) Identification of Other domains Involved in K+ Channel Subunit Multimerization, 3) Characterization of the Subfamily Dependent Differences in the T1 Domain and Testing Whether T1 Subfamily Specificity is Dominant, and 4) Examination of the Role of T1 Domain Compatibility in the Formation of Functional K+ Channel Proteins. The experiments to accomplish these Specific Aims rely exclusively on techniques or methods that are presented in the Preliminary Results or are being used currently in the lab. The general approach is to clone subfragments of different K+ channel subunit proteins that are thought to encode T1 domains, and to test for their function and subfamily specificity. A series of mutagenesis experiments will identify specific regions and amino acid residues that are critical for T1 domain tetramerization and subfamily specificity. Finally, a set of chimeric K+ channel subunit proteins will be created, with swapped T1 domains, to test for the importance of T1 in determining subunit protein interaction and formation of homomultimeric and heteromultimeric K+ channels. Because K+ channel proteins are an increasingly important target for drugs to treat asthma, high blood pressure, diabetes, multiple sclerosis, epilepsy and potentially a host of other disorders, these experiments will benefit health research by providing critical information about the events and domains of K+ channel subunit proteins that are involved in forming the ion channel.
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1993 — 2000 |
Pfaffinger, Paul |
P30Activity Code Description: To support shared resources and facilities for categorical research by a number of investigators from different disciplines who provide a multidisciplinary approach to a joint research effort or from the same discipline who focus on a common research problem. The core grant is integrated with the center's component projects or program projects, though funded independently from them. This support, by providing more accessible resources, is expected to assure a greater productivity than from the separate projects and program projects. R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Functional Domains of Potassium Channel Subunit Proteins @ Baylor College of Medicine
DESCRIPTION: K+ channels are an important target for drugs to treat a variety of disorders, including high blood pressure, diabetes, asthma, multiple scleroisis, and others. In addition, the genetic disease, Periodic Ataxia and certain forms of Long Q-T syndrome, have been linked to specific point mutations in K+ channel proteins. Despite the importance these channels have in maintaining normal physiology, and their net therapeutic importance, relatively little is known about how K+ channel assembly by the N-terminal T1 domain. The experiments the investigator will perform in this proposal are motivated by the progress he has achieved in understanding the assembly mechanisms of voltage gated K channels from the original proposal. In this new project, the investigator will determine the crystallographic structure of the T1 domain protein in collaboration with Dr. Senyon Choe, confirm the structural model in isolated T1 domain proteins and full length channels, identify critical residues and interactions for K channel assembly, and characterize the alterations in channel assembly identity produced by T1 domain mutations. The long term goal is to understand how information needed to encode the precise electrophysiological properties of the nervous system are encoded by structural differences in the T1 domain of K channel subunit proteins. This goal will be addressed by experiments that will satisfy the following Specific Aims: 1) Determination of the structure of the tetrameric T1 domain and identification of functionally significant regions of the T1 domain protein; 2) analysis of residues that are functionally important for T1 domain tetramerization and confirmation of the T1 domain structure; and 3) analysis of the T1 domain in the full length channel subunit protein and its role in controlling channel protein assembly. The general approach in these studies is to integrate structural information with functional information on wild type and mutant T1 domain assembly in isolated protein preparations, and for full length subunit protein assembly, and intact cells. By careful characterization of the changes in assembly properties caused by these individual mutations, the investigators can dissect the functional roles for specific residues and residue interactions in producing the ordered assembly of K channel subunit protein in the nervous system.
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1999 — 2002 |
Pfaffinger, Paul |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Characterize Molecular Properties of Mammalian Neuronal Dendritic K Channel @ Baylor College of Medicine
neuroregulation; potassium channel; dendrites; hippocampus; protein structure function; pyramidal cells; protein protein interaction; protein sequence; protein kinase A; phosphorylation; protein kinase C; mitogen activated protein kinase; voltage gated channel; yeast two hybrid system; site directed mutagenesis; genetically modified animals; laboratory mouse; Xenopus; laboratory rabbit; laboratory rat; voltage /patch clamp; immunofluorescence technique; fluorescence microscopy; antisense nucleic acid;
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2004 — 2006 |
Pfaffinger, Paul |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Molecular Properties of Neuronal Dendritic K+ Channels @ University of Texas Austin
Our general working hypothesis is that the functional properties of native voltage-gated potassium channels arise through the interactions of different auxiliary proteins with the core pore forming alpha subunits. This hypothesis emphasizes the importance of protein-protein interactions in the formation, trafficking, and regulation of neuronal electrical properties. In this proposal we will be testing the specific hypothesis that Kv4 alpha subunits, also called Shal type Kv channels, assemble with auxiliary KChlP proteins and DPP proteins to form a macromolecular protein complex that is trafficked to the dendrites in CA1 pyramidal neurons to form a functionally important A current channel. Our studies will focus on the analysis of changes in A current function in native neurons in response to molecular genetic manipulations in expression level for different subunits in native neurons. Next, we will examine the functional effects of introducing mutant version of the important subunits proteins that either lack the ability to perform specific protein-protein interactions or are unable to be modified by particular post-translational modifications. By characterizing the changes in function at several different levels, we will determine the specific roles of different subunit proteins, functional motifs, and post-translational modifications in forming the A current phenotypes of CA1 pyramidal neurons. These studies will have a general impact on our understanding of the molecular control of neuronal electrical phenotypes as well as the approaches we will take in understand the functional importance of other ion channels and auxiliary subunit proteins. Our studies will have a general impact on our understanding of the regulation of excitability that will impact many areas of health related research including epilepsy, heart disease, asthma, and AIzheimer's.
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2007 — 2008 |
Pfaffinger, Paul |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Molecular Properties of Mammalian Neuronal Dendritic K+ Channels @ University of Texas Austin
Our general working hypothesis is that the functional properties of native voltage-gated potassium channels arise through the interactions of different auxiliary proteins with the core pore forming alpha subunits. This hypothesis emphasizes the importance of protein-protein interactions in the formation, trafficking, and regulation of neuronal electrical properties. In this proposal we will be testing the specific hypothesis that Kv4 alpha subunits, also called Shal type Kv channels, assemble with auxiliary KChlP proteins and DPP proteins to form a macromolecular protein complex that is trafficked to the dendrites in CA1 pyramidal neurons to form a functionally important A current channel. Our studies will focus on the analysis of changes in A current function in native neurons in response to molecular genetic manipulations in expression level for different subunits in native neurons. Next, we will examine the functional effects of introducing mutant version of the important subunits proteins that either lack the ability to perform specific protein-protein interactions or are unable to be modified by particular post-translational modifications. By characterizing the changes in function at several different levels, we will determine the specific roles of different subunit proteins, functional motifs, and post-translational modifications in forming the A current phenotypes of CA1 pyramidal neurons. These studies will have a general impact on our understanding of the molecular control of neuronal electrical phenotypes as well as the approaches we will take in understand the functional importance of other ion channels and auxiliary subunit proteins. Our studies will have a general impact on our understanding of the regulation of excitability that will impact many areas of health related research including epilepsy, heart disease, asthma, and AIzheimer's.
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0.903 |
2008 — 2009 |
Pfaffinger, Paul |
P41Activity Code Description: Undocumented code - click on the grant title for more information. |
Protein-Protein Interactions in Voltage-Gated Potassium Channel Assembly and Ga @ Carnegie-Mellon University
Acceleration; Binding Sites; Biochemistry; Biophysics; CRISP; Chemistry, Biological; Combining Site; Complex; Computer Retrieval of Information on Scientific Projects Database; Family member; Funding; Grant; Institution; Investigators; Ion Channel; Ionic Channels; KCNA2 potassium channel; KCND2 channel; Kv1.2 gene product; Kv1.2 potassium channel; Kv1.2'channel; Kv4.2 channel; Membrane Channels; Models, Structural; Molecular Dynamics Simulation; N-terminal; NH2-terminal; NIH; National Institutes of Health; National Institutes of Health (U.S.); Peptides; Property; Property, LOINC Axis 2; Protein Binding; Protein Subunits; Proteins; Reactive Site; Recovery; Research; Research Personnel; Research Resources; Researchers; Resources; Role; Source; Structural Models; Structure; TM Domain; Transmembrane Domain; Transmembrane Region; United States National Institutes of Health; Voltage-Gated K+ Channels; Voltage-Gated Potassium Channel; gene product; insight; interfacial; molecular dynamics; protein protein interaction; social role; structural biology
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0.907 |
2010 — 2013 |
Pfaffinger, Paul |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
A Current Regulation by Dipeptidyl Peptidase-Like Proteins @ Baylor College of Medicine
DESCRIPTION (provided by applicant): A Current Regulation by Dipeptidyl Peptidase-Like Proteins Dipeptidyl peptidase like (DPL) proteins, DPP6 and DPP10, regulate Kv4 channel expression and functional properties and are essential components of the native neuronal ISA, somatodendritic A current along with Kv4 alpha subunits and KChIP auxiliary subunits. Without DPL expression, ISA is severely disrupted with reduced current expression and abnormal activation and inactivation properties. In this project we will test the hypothesis that specific conserved functional domains in the first two exons of DPL genes regulate the interaction of DPL proteins with Kv4 channels proteins and determine the functional properties of the channel complex. These studies will provide important new information about the molecular mechanisms that regulate the functional properties of neurons and likely will be important for our understanding of the molecular mechanisms underlying disease processes such as ALS, autism spectrum disorder, asthma, and other regulatory pathways that DPL proteins have been implicated in. In this project we will address the following aims to better understand the molecular mechanisms involved in the regulation of A currents by DPL proteins. Aim 1: Test the hypothesis that DPP6a and DPP10a accelerate inactivation using a novel N-terminal motif that shares a common underlying molecular mechanism with other N-type inactivation domains. Aim 2: Test the Hypothesis that multiple intermediate states are experienced during N-type inactivation by DPP6a and DPP10a. Aim 3: Test the hypothesis that specific DPL residues in transmembrane and peri-transmembrane region modulate Kv4 channel activation gating.
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2019 |
Arenkiel, Benjamin R (co-PI) [⬀] Pfaffinger, Paul |
U01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Multilevel Analysis of Neuronal Computations Underlying the Robust Encoding of Sensory Information in the Mammalian Olfactory System @ Baylor College of Medicine
A key problem in neuroscience is to uncover fundamental principles and algorithms that allow neuronal networks to perform the complex calculations that underlie normal behavior effectively and efficiently. Our goal in this project is to develop a powerful team approach to understand the mechanisms and principles underlying the processing of odorant information by the olfactory system. In Project 1, we will develop the experimental and computational methods to characterize the processing of olfactory information by the olfactory bulb, and to examine how that processing changes with learning or changes in neuronal excitability. In Project 2, we will develop real time approaches to the analyses developed in Project 1 to improve our ability to test hypotheses of information encoding in the olfactory bulb, and the roles of specific neurons in this encoding. In Project 3, we will perform neural network modeling of the olfactory bulb circuit in a way that allows rapid and global optimization of parameters that have biophysical relevance for understanding bulb circuitry function. In Project 4, we will develop dual in vivo imaging of cortex and olfactory bulb to better understand the transfer of olfactory information to the cortex and how this changes with learning. In Project 5, we will examine the cortical feedback to the bulb to understand how this feedback helps shape olfactory odorant responses and direct the appropriate changes in bulb circuitry during learning that will preserve odor information. The technical innovation of this proposal is driven by a multilevel experimental approach that leverages expertise from an investigative team with diverse backgrounds. We expect this approach to uncover valuable insights into principles of neural circuit organization and algorithmic function that underlie olfactory system function and plasticity. Our studies will have a broad impact on our understanding of how neuronal circuits implement effective algorithms, and will likely provide important insights into the function of other circuits in the central nervous system. In addition, our work may reveal previously unrecognized computational algorithms that will have a broad impact on computer science.
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