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The molecular basis of normal development, regeneration and myelin-related neuropathies in peripheral nerves

Objective

To understand the mechanisms that regulate normal nerve development at the molecular level. It is believed that this provides an essential framework for understanding Schwann cell malfunction in debilitating neuropathies and the processes that underlie nerve repair. Methodologically, the approach is characterised by a very close integration of advanced molecular biology techniques with cell biology.

To define genes that are important in Schwann cell development and nerve regeneration, and to directly relate such genes to normal and diseased human Schwann cells. Transcription factors that hold a key position in the regulation of Schwann cell development are beginning to emerge, one of the most important of these being the POU domain protein Oct. 6.

To prepare a detailed analysis of Oct 6 function with the aim of understanding its pivotal role in Schwann cell differentiation, particularly myelin formation.

To generate an animal model of the most common hereditary human neuropathy Charcot-Marie-Tooth (CMT) disease.

We aim to understand the mechanisms that regulate normal nerve development at the molecular level. We believe that this provides an essential framework for understanding Schwann cell malfunction in debilitating neuropathies and the processes that underlie nerve repair. Methodologically, our approach is characterized by a very close integration of advanced molecular biology techniques with cell biology. A major emphasis will be on defining genes that are important in Schwann cell development and nerve regeneration, and to directly relate such genes to normal and diseased human Schwann cells. Transcription factors that hold a key position in the regulation of Schwann cell development are beginning to emerge, one of the most important of these being the POU domain protein Oct 6. Another priority area will be a detailed analysis of Oct 6 function with the aim of understanding its pivotal role in Schwann cell differentiation, particularly myelin formation. A third main objective is to generate an animal model of the most common hereditary human neuropathy Charcot-Marie-Tooth (CMT) disease.

The project is divided into six Tasks
I) Identification and functional characterization of qenes that are differentially regulated in Schwann cell/Peripheral nerve development and regeneration. Differential display techniques will be used to complete mRNA from two different stages of development, or from normal and transacted nerves.

Genes that show differential expression will be subcloned, sequenced and their expression analysed by in situ hybridisation. Functional characterization of kegenes will be carried out in vitro following transfection or microinjection, or in transgenic mice, and expression in normal and pathological human nerves will be examined;
(II) Defininq the role of Oct 6 in Schwann cell development: a cell biological approach. Schwann cells and Schwann cell precursors from an Oct 6-/- mouse, which show a striking inhibition of myelination, will be analysed in well defined culture systems to uncover abnormalities in survival, DNA synthesis and differentiation;
(III) Identification of aenes that are regulated by the Oct 6 protein. To reveal Schwann cell genes that are regulated by Oct 6, differential display methods will be used to compare mRNA from normal and Oct 6-/- nerves, and a large range of known Schwann cell related genes/proteins will be compared in normal and -/- nerves;
(IV) Identification of cis-acting DNA elements in the Oct 6 promoter and of aenes/Proteins that regulate Oct 6. Transgenic mice, DNAse 1 foot printing and related methods and cell culture will be used to identify regulatory sequences and proteins that bind to and/or regulate them;
(V) Oct 6 binding partners. We will use the Yeast-2-hybrid system to identify proteins that bind to Oct 6;
(VI) Refinement of a PO promoter-based transaenic cassette and creation of mous models of CMTlb neuropathies. The PO promoter and transgenic mice will be used to study the role of the Pax-3transcription factor in myelination. To establish a mouse model of CMTlb, we will use similar experiments to test whether expression of Pc with targeted, CMTlb-like mutations leads to CMTlb-like neuropathies. 04

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

UNIVERSITY COLLEGE LONDON
Address
Gower Street
London
United Kingdom

Participants (3)

Erasmus Universiteit Rotterdam
Netherlands
Address
50,Dr. Molewaterplein
3000 DR Rotterdam
Fondazione Centro San Raffaele del Monte Tabor - Istituto di Ricovero e Cura a Carattere Scientifico
Italy
Address
Via Olgettina
20132 Milano
Heinrich-Heine-Universität Düsseldorf
Germany
Address
5,Moorenstrasse
40225 Düsseldorf