Posttranslational modification of CLN3 protein and its possible functional implication.
The CLN3 gene associated with Batten disease and encoding a novel protein of a predicted 438 amino acids was cloned in 1995 by the International Batten Disease Consortium. The function of CLN3 protein remains unknown. Computer-based analysis predicted that CLN3 may contain several posttranslational modifications. Thus, to study the posttranslational modification of CLN3 protein, we have expressed a full-length CLN3 protein as a C-terminal fusion with green fluorescent protein of the jellyfish Aequerea victoria in a Chinese hamster ovary cell line. Previously, we have shown that CLN3 is a glycosylated protein from lysosomal compartment, and now, by using in vivo labeling with 32P, detection with anti-phosphoamino acid antibodies, and phosphoamino acid analysis, we demonstrate that CLN3 is a phosphorylated protein. We demonstrate that CLN3 protein does not undergo mannose 6-phosphate modification and that it is a membrane protein. Furthermore, we show that the level of CLN3 protein phosphorylation may be modulated by several protein kinases and phosphatases activators or inhibitors.
Michalewski
M P
MP
Department of Pathological Neurobiology, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York 10314, USA.
Kaczmarski
W
W
Golabek
A A
AA
Kida
E
E
Kaczmarski
A
A
Wisniewski
K E
KE
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D016213:Cyclins; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000378:metabolism; D049452:Green Fluorescent Proteins; D015151:Immunoblotting; D008164:Luminescent Proteins / Q000378:metabolism; D008360:Mannosephosphates / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D018832:Molecular Chaperones / Q000378:metabolism*; D019606:Phosphoamino Acids / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D011499:Protein Processing, Post-Translational; D011868:Radioisotopes / Q000378:metabolism; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine / Q000378:metabolism; D013912:Threonine / Q000378:metabolism
C079895:CLN3 protein, S cerevisiae; C105199:CLN3 protein, human; D016213:Cyclins; D004791:Enzyme Inhibitors; D008164:Luminescent Proteins; D008360:Mannosephosphates; D008562:Membrane Glycoproteins; D018832:Molecular Chaperones; D019606:Phosphoamino Acids; D011868:Radioisotopes; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins; D013912:Threonine; C027693:mannose-6-phosphate; D012694:Serine; D000469:Alkaline Phosphatase; D006026:Glycoside Hydrolases