Posttranslational modification of CLN3 protein and its possible functionalimplication

The CLN3 gene associated with Batten disease and encoding a novel protein o f a predicted 438 amino acids was cloned in 1995 by the International Batte n Disease Consortium, The function of CLN3 protein remains unknown. Compute r-based analysis predicted that CLN3 may contain several posttranslational modifications, Thus, to study the posttranslational modification of CLN3 pr otein, we have expressed a full-length CLN3 protein as a C-terminal fusion with green fluorescent protein of the jellyfish Aequerea victoria in a Chin ese hamster ovary cell line. Previously, we have shown that CLN3 is a glyco sylated protein from lysosomal compartment, and now, by using in vivo label ing with P-32, detection with anti-phosphoamino acid antibodies, and phosph oamino acid analysis, we demonstrate that CLN3 is a phosphorylated protein. We demonstrate that CLN3 protein does not undergo mannose 6-phosphate modi fication and that it is a membrane protein, Furthermore, we show that the l evel of CLN3 protein phosphorylation may be modulated by several protein ki nases and phosphatases activators or inhibitors. (C) 1999 Academic Press.