Nerve growth factor (NGF) induces increase in multi-ubiquitin chains and concomitant decrease in free ubiquitin in nuclei of PC12h.
Changes in Ubiquitin-immunoreactivity after nerve growth factor (NGF) treatment were investigated in PC12h cells. Ubiquitin-immunoreactivity was increased in the nucleus of NGF-treated cells. The quantitative analysis revealed that, after 7 days of NGF treatment, almost 20% of cells had ubiquitin-immunoreactive nuclei and the frequency was increased thereafter. Levels of free ubiquitin and multi-ubiquitin chains were measured by radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA), respectively. Measurements were carried out for four subcellular fractions: urea- and water-soluble extracts of nuclei and cytoplasm. Decrease in free ubiquitin was observed in water-soluble cytoplasmic extracts of NGF-treated cells, though increase in multi-ubiquitin chains in the same fraction was not observed. As for nuclei, increase in multi-ubiquitin chains and concomitant decrease in free ubiquitin were found in the water-soluble extracts after NGF treatment. Levels of multi-ubiquitin chains did not change in urea-soluble cytoplasmic extracts as well as nuclear urea-soluble ones after NGF treatment. These results indicated that multi-ubiquitination of nuclear proteins is increased during NGF-induced neuronal differentiation of PC12h cells.
Ohtani-Kaneko
R
R
Department of Anatomy, St. Marianna University School of Medicine, Kanagawa, Japan.
Asahara
M
M
Takada
K
K
Kanda
T
T
Iigo
M
M
Hara
M
M
Yokosawa
H
H
Ohkawa
K
K
Hirata
K
K
D000818:Animals; D002478:Cells, Cultured / Q000378:metabolism; D007150:Immunohistochemistry; D009414:Nerve Growth Factors / Q000494:pharmacology*; D016716:PC12 Cells / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D014452:Ubiquitins / Q000378:metabolism*
D009414:Nerve Growth Factors; D014452:Ubiquitins