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The change that accompanies the prion protein from its normal
to abnormal Isoform involves a considerable increase in the beta sheet
content.
Methods like CD
spectroscopy, Urea gradient electrophoresis can be used to study the transition between the normal
and abnormal isoforms.
If one can manipulate the aromatic amino acids like tryptophan, can Fluorescence spectroscopy be employed?
Site Directed mutagenesis studies can also be employed to understand which residues are
crucial to the change in structure.
Data obtained from these studies will help us also determine
which secondary structures are important for the changes that accompany the
prion protein.
SIGNAL TRANSDUCTION
Prions are believed to be involved in Copper ion
concentration maintenance. Recently it has also been shown that a loss of
LPS induced Nitric Oxide Production and Nitric oxide synthase
expression occurs in scrapie infected cells.
Approach:
1. Make luciferase
constructs of the Prion Protein promoter.. AP-1,
AP-2, inverted CCAATmotif, and Sp1 sites are also
present in the promoter region of the Prion protein.
2. Sp1 is associated with HDACs. Hence chromatin remodeling might affect levels
of gene expression. Inhibition of HDACs can be performed
using various HDAC inhibitors and determine luciferase
activity.
3. Cotransfect
cells with different types of potential regulators of Prion Protein gene
expression e.g. CREB. One might also be able to grow cells in presence of
various hormones like Nerve Growth Factor, Retinoic acid and drugs known to
inhibit neural growth.
4. For each of these assays the
relative levels of prion protein expression, one might consider performing
Northern Blots and Western blots.
Reference:
Ana L.B. Cabral, Kil S. Lee and Vilma R. Martins – Journal of Biological
Chemistry – Vol 277 No.7 Feb 15th
; 5675-5682
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