PATHOGENESIS OF BATTEN DISEASE

University Of Chicago, Chicago IL

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Abstract

We propose to continue studies on the pathogenesis of Battens disease by focusing on the Infantile form of Batten Disease (CLN 1) and the deficient enzyme palmitoyl:protein thioesterase (PPT 1). We will overexpress PFF 1 in human neuroblastoma LA-N-5 cells, PC 12 pheochromocytoma cells, embryonic chick neurons, and non-neural CHO cells using inducible promoters, fluorescent tags and specific sequences to direct expression to discrete subcellular compartments such as mitochondria. We will determine how PPT1 is able to regulate the level of protein palmitoylation in detergent-resistant microdomains (Rafts) and hence the level of activity of Akt and the phosphatase PTEN in order to protect against cell death. We will verify the presence of PPT1 activity in Rafts and the relationship between raft and lysosomal PPT1. We will use gene array technology to determine which cell death-associated genes are turned on/off when PPT1 is overexpressed and in situ hybridization to follow the spatio-temporal expression of PPT1 in the developing nervous system. In order to model the deficiency of PPT1 we will synthesize specific, potent inhibitors of PPT1 based on AcG-palmitoyl diamino propionate-VKIKK (DAP1) and ketoamide analogs of the palmitoyl-cysteine thioester linkage. We will characterize these inhibitors for optimum inhibition of PPT1 and cell uptake. We will determine if the cell death they cause results from inactivation of Akt and use them to better understand the role of PPT1 in Rafts. Our broad, long-term objectives are to understand the reason for the early death of cortical neurons in CLN 1 and how this might be remedied.