Nuclear substrates of protein kinase C.
Starting from the finding that, for neuronal cells, the nuclear-membrane-associated protein kinase C (PKC) is the so-called 'membrane inserted', constitutively active form, we attempted to identify substrates of this nuclear PKC. For this purpose, nuclear membranes and other subcellular fractions were prepared from bovine brain, and in-vitro phosphorylation was performed. Several nuclear membrane proteins were found, the phosphorylation of which was inhibited by specific PKC inhibitors and effectively catalyzed by added PKC. Combining the methods of two-dimensional gel electrophoresis, in-situ digestion, reverse-phase HPLC and microsequencing, two of these nuclear PKC substrates were identified; the known PKC substrate Lamin B2, which serves as a control of the approach and the nucleolar protein B23. Our data suggest, that, for B23, Ser225 is a site of phosphorylation by PKC.
Beckmann
R
R
Institut für Biochemie, Freie Universität Berlin, Federal Republic of Germany.
Buchner
K
K
Jungblut
P R
PR
Eckerskorn
C
C
Weise
C
C
Hilbert
R
R
Hucho
F
F
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002467:Cell Nucleus / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D007425:Intracellular Membranes / Q000201:enzymology*; D034921:Lamin Type B; D034882:Lamins; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D000090243:Nucleophosmin; D010449:Peptide Mapping; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013379:Substrate Specificity; D014357:Trypsin
D034921:Lamin Type B; D034882:Lamins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D000090243:Nucleophosmin; D011493:Protein Kinase C; D014357:Trypsin