2001 — 2005 |
Bermingham, John Rutledge |
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. |
Genetic Control of Schwann Cell Myelination @ Mc Laughlin Research Institute
DESCRIPTION (From the Applicant's Abstract): In spite of the medical importance of myelin, as evidenced by the debilitating effects of demyelinating diseases, little is known about the genetic regulatory mechanisms that control myelin synthesis. Axonal contact triggers myelination through as yet unknown signal(s), many of which can be mimicked in Schwann cells by treatment with the adenylyl cyclase activator forskolin. The POU domain transcription factor Oct-6 (also known as Tst-1 or SCIP) is one of the genes that is induced by forskolin. The phenotypes of mice that lack Oct-6 reveal that it plays a critical role in controlling the timing and rate of peripheral myelination, but the mechanisms by which it does so are unknown. A screen for additional forskolin-regulated genes produced clones for the ERK2-specific phosphatases MKP-3, and the sphingosine-1 phosphate (S1P) receptor edg-3. To better understand the role of these signal transduction molecules in Schwann cell differentiation, the first Specific Aim of this application will study their expression patterns in Schwann cells, and their ability to activate differentiation markers such as Oct-6 in sciatic nerve, of which the functions and/or identifies of five are unknown. The second Specific Aim of this application is to characterize these Oct-6 induced genes genes further. The two known genes that are activated by Oct-6, the cytoplasmic LIM domain protein CRP2, and the fatty acid transport protein myelin P2, suggest cytoskeletal rearrangement or fatty acid transport as rate-limiting steps in myelination. Both genes possess putative Oct-6 binding sites in their promoters. The hypothesis that Oct-6 directly regulates the expression of these genes will be tested in the third Specific Aim of this application. In the fourth Specific Aim, it is proposed to study postnatal functions of Oct-6 in peripheral myelination using mice that lack the gene only in Schwann cells.
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0.988 |
2007 — 2011 |
Bermingham, John Rutledge |
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. |
Genetic Control of Schwann Cell Differentiation @ Mc Laughlin Research Institute
[unreadable] DESCRIPTION (provided by applicant): Project Summary: Myelin is a crucial component of vertebrate nervous systems. Both regenerative therapies and treatment of demyelinating diseases will be facilitated by knowledge of the mechanisms that control its synthesis. These diseases are frequent, and account for considerable morbidity and mortality. However, the regulation of myelination and remyelination is poorly understood. In the PNS, Schwann cells require axonal contact for proliferation, survival, and for the expression of the transcription factor Oct-6, which is required for timely myelination. In claw paw mutant mice, both the segregation of axon fascicles by Schwann cells and their expression of Oct-6 downstream effector genes is delayed, even though Oct-6 is activated normally. Recently, we have identified the genetic defect underlying the claw paw mutation as an insertion in the Lgi4 (Leucine-rich glioma inactivated-4) gene. This gene is expressed by Schwann cells, and encodes a secreted protein of unknown function. Thus Schwann cells secrete a protein that they require for axon segregation, activation of Oct-6 target genes, and myelination. We hypothesize that Lgi4 triggers or modifies axonal or extracellular matrix signal(s), thereby regulating Schwann cell responses to these cues. We propose the following aims to determine how Lgi4 functions in Schwann cell development: 1) Test the hypothesis that Schwann cell differentiation is modulated by the level of Lgi4 expression. 2) Test the hypothesis that Lgi4 controls the expression of essential Schwann cell genes. 3) Test the hypothesis that Lgi4 acts through the Akt/PKB signaling pathway. The experiments summarized above focus on identifying Schwann cell signaling pathways that respond to Lgi4-mediated extracellular signaling; in doing so, we will elucidate the genetic regulatory pathways that result in the formation of peripheral myelin, thereby providing the foundation for new therapies to treat myelination disorders. [unreadable] [unreadable] Relevance: Diseases that affect myelin, such as multiple sclerosis in the central nervous system, and inherited demyelinating neuropathies in the peripheral nervous system, are both debilitating and frequent. By understanding how the Lgi4 protein controls Schwann cell development, we will establish new avenues for the treatment of myelin disorders and nerve injury. [unreadable] [unreadable] [unreadable]
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0.988 |
2010 |
Bermingham, John Rutledge |
S10Activity Code Description: To make available to institutions with a high concentration of NIH extramural research awards, research instruments which will be used on a shared basis. |
Ge Typhoon 9410 Variable Mode Imager @ Mc Laughlin Research Institute
DESCRIPTION (provided by applicant): Imaging and quantifying labeled molecules is a critical component of many modern biomedical research techniques. For the last 11 years we have imaged the results of Southern and northern hybridizations, and RNAse protections on a Molecular Dynamics Phosophorimager SI. The results must be analyzed using software on an old Macintosh computer that runs obsolete System 8 software. Should the phosphorimager or computer fail, they cannot be serviced. After 11 years of use, the phosphorimaging screens have deteriorated past their useful lifetime. The McLaughlin Research Institute is an independent non-profit research institute in Great Falls, Montana, three hours'drive away from other facilities with phosphor imaging capability. Although Southern and northern hybridization experiments continue to rely on radio isotopic labeling, fluorescent labeling techniques continue to advance, with new applications in imaging nucleic acids and proteins. We need to replace our phosphor imager with one that is capable of imaging and quantifying both radioactive and fluorescently labeled molecules. We are requesting the General Electric Typhoon 9410 Variable Mode Imager. In addition to functioning as a traditional phosphor imager, the Typhoon 9410 can detect red, green and blue fluorescence, permitting multiple nucleic acids and proteins to be detected simultaneously. This fluorescent capability will permit us perform multiplex PCR reactions for genotyping, as well as detection of proteins transferred to membranes from acrylamide gels (western blots) or from tissue sections (histoblots). The Typhoon 9410 has 10 micron resolution, a particularly useful feature for histoblots. The updated phosphor imaging capabilities that are provided by this equipment will permit us to more efficiently and reliably perform experiments using radio labeled nucleic acids, but will expand our capability to perform state-of-the-art techniques using labeled proteins as well. Public Health Relevance: To image and quantify radiolabelled molecules that are essential for our research, we have relied on a Molecular Dynamics Phosophorimager SI, whose capabilities have deteriorated considerably over the last 11 years. Although radioisotope labeling remains a useful research technique, fluorescent labeling techniques continue to advance. We need to replace our Phosphor imager with one that is capable of imaging and quantifying both radioactive and fluorescently Labeled molecules to advance our research, which is relevant to neurodegenerative diseases, Parkinson's disease and myelin diseases.
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0.988 |