Evidence for phosphorylation of CLN3 protein associated with Batten disease

Recently, the CLN3 gene associated with Batten disease (juvenile neuronal c eroid lipofuscinosis, JNCL), a recessively inherited, progressive, neurodeg enerative disorder of childhood, has been identified. The CLN3 gene encodes a novel protein (battenin) of a predicted 438 amino acids containing sever al potential posttranslational modifications. We have expressed a full-leng th CLN3 protein as a C-terminal fusion with green fluorescent protein (GFP) to evaluate whether CLN3 protein is phosphorylated, By using in vivo label ing with (32)p, detection with anti-phosphoamino acid antibodies, and phosp hoamino acid analysis, we demonstrate that the CLN3 protein is phosphorylat ed on both serine and threonine residues. We also demonstrate that CLN3 pro tein is not modified by mannose 6-phosphate. Furthermore, we show that phos phorylation of CLN3 protein is carried out by protein kinase A (cAMP-depend ent protein kinase, PKA), protein kinase G (cGMP-dependent protein kinase, PKG), and casein kinase LT and that it is enhanced by inhibition of protein phosphatase I (PP 1) or protein phosphatase 2A (PP 2A). (C) 1998 Academic Press.