PMID- 12130654
OWN - NLM
STAT- MEDLINE
DCOM- 20021113
LR  - 20210209
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 277
IP  - 39
DP  - 2002 Sep 27
TI  - In vivo and in vitro phosphorylation at Ser-493 in the glutamate (E)-segment of 
      neurofilament-H subunit by glycogen synthase kinase 3beta.
PG  - 36032-9
AB  - Neurofilament (NF), a major neuronal intermediate filament, is composed of three 
      subunits, NF-L, NF-M, and NF-H. All three subunits contain a well conserved 
      glutamate (E)-rich region called "E-segment" in the N terminus of the tail 
      region. Although the E-segments of NF-L and NF-M are phosphorylated by casein 
      kinases, it has not been observed in NF-H. Using mass spectrometric analysis, we 
      identified phosphorylation of the E-segment of NF-H, prepared from rat spinal 
      cords, at Ser-493 and Ser-501 in the Ser-Pro sequences. The E-segment kinase was 
      isolated from rat brain extract using column chromatography and identified as 
      glycogen synthase kinase (GSK) 3beta. GSK3beta was shown to phosphorylate at 
      Ser-493 in vitro by phosphopeptide mapping and site-directed mutagenesis, and in 
      vivo in HEK293 cells using the phospho-Ser-493 antibody, but did not 
      phosphorylate Ser-501. GSK3beta preferred Ser-493 to the KSP-repeated sequences 
      for phosphorylation sites in the NF-H tail domain. Moreover, Ser-493 was a better 
      phosphorylation site for GSK3beta than other proline-directed protein kinases, 
      Cdk5/p35 and ERK. GSK3beta in the spinal cord extract was associated with NF 
      cytoskeletons. Taken together, we concluded that Ser-493 in the E-segment of NF-H 
      is phosphorylated by GSK3beta in rat spinal cords.
FAU - Sasaki, Takahiro
AU  - Sasaki T
AD  - Department of Biological Sciences, Graduate School of Science, Tokyo Metropolitan 
      University, Minami-ohsawa, Hachiohji, Tokyo 192-0397, Japan. 
      sasaki-takahiro@c.metro-u.ac.jp
FAU - Taoka, Masato
AU  - Taoka M
FAU - Ishiguro, Koichi
AU  - Ishiguro K
FAU - Uchida, Atsuko
AU  - Uchida A
FAU - Saito, Taro
AU  - Saito T
FAU - Isobe, Toshiaki
AU  - Isobe T
FAU - Hisanaga, Shin-Ichi
AU  - Hisanaga S
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20020718
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - 0 (Neurofilament Proteins)
RN  - 108688-71-7 (neurofilament protein H)
RN  - 3KX376GY7L (Glutamic Acid)
RN  - 452VLY9402 (Serine)
RN  - 9DLQ4CIU6V (Proline)
RN  - EC 2.7.11.1 (Cyclin-Dependent Kinase 5)
RN  - EC 2.7.11.1 (GSK3B protein, human)
RN  - EC 2.7.11.1 (Glycogen Synthase Kinase 3 beta)
RN  - EC 2.7.11.1 (Gsk3b protein, rat)
RN  - EC 2.7.11.22 (CDK5 protein, human)
RN  - EC 2.7.11.22 (Cdk5 protein, rat)
RN  - EC 2.7.11.22 (Cyclin-Dependent Kinases)
RN  - EC 2.7.11.24 (Mitogen-Activated Protein Kinases)
RN  - EC 2.7.11.26 (Glycogen Synthase Kinase 3)
RN  - EC 3.4.- (Endopeptidases)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Binding Sites
MH  - Blotting, Western
MH  - Brain/metabolism
MH  - Cell Line
MH  - Cloning, Molecular
MH  - Cyclin-Dependent Kinase 5
MH  - Cyclin-Dependent Kinases/metabolism
MH  - Cytoskeleton/metabolism
MH  - Electrophoresis, Polyacrylamide Gel
MH  - Endopeptidases/metabolism
MH  - Glutamic Acid/metabolism
MH  - Glycogen Synthase Kinase 3/*metabolism
MH  - Glycogen Synthase Kinase 3 beta
MH  - Humans
MH  - Mitogen-Activated Protein Kinases/metabolism
MH  - Molecular Sequence Data
MH  - Neurofilament Proteins/*chemistry
MH  - Phosphorylation
MH  - Precipitin Tests
MH  - Proline/chemistry
MH  - Protein Binding
MH  - Protein Structure, Tertiary
MH  - Rats
MH  - Serine/*chemistry/metabolism
MH  - Spinal Cord/metabolism
MH  - Time Factors
EDAT- 2002/07/20 10:00
MHDA- 2002/11/26 04:00
CRDT- 2002/07/20 10:00
PHST- 2002/07/20 10:00 [pubmed]
PHST- 2002/11/26 04:00 [medline]
PHST- 2002/07/20 10:00 [entrez]
AID - S0021-9258(18)36568-2 [pii]
AID - 10.1074/jbc.M206674200 [doi]
PST - ppublish
SO  - J Biol Chem. 2002 Sep 27;277(39):36032-9. doi: 10.1074/jbc.M206674200. Epub 2002 
      Jul 18.