PMID- 20169205
OWN - NLM
STAT- MEDLINE
DCOM- 20100930
LR  - 20211020
IS  - 1932-6203 (Electronic)
IS  - 1932-6203 (Linking)
VI  - 5
IP  - 2
DP  - 2010 Feb 12
TI  - Raptor is phosphorylated by cdc2 during mitosis.
PG  - e9197
LID - 10.1371/journal.pone.0009197 [doi]
LID - e9197
AB  - BACKGROUND: The appropriate control of mitotic entry and exit is reliant on a 
      series of interlocking signaling events that coordinately drive the biological 
      processes required for accurate cell division. Overlaid onto these signals that 
      promote orchestrated cell division are checkpoints that ensure appropriate 
      mitotic spindle formation, a lack of DNA damage, kinetochore attachment, and that 
      each daughter cell has the appropriate complement of DNA. We recently discovered 
      that AMP-activated protein kinase (AMPK) modulates the G2/M phase of cell cycle 
      progression in part through its suppression of mammalian target of rapamycin 
      (mTOR) signaling. AMPK directly phosphorylates the critical mTOR binding partner 
      raptor inhibiting mTORC1 (mTOR-raptor rapamycin sensitive mTOR kinase complex 1). 
      As mTOR has been previously tied to mitotic control, we examined further how 
      raptor may contribute to this process. METHODOLOGY/PRINCIPAL FINDINGS: We have 
      discovered that raptor becomes highly phosphorylated in cells in mitosis. 
      Utilizing tandem mass spectrometry, we identified a number of novel 
      phosphorylation sites in raptor, and using phospho-specific antibodies 
      demonstrated that raptor becomes phosphorylated on 
      phospho-serine/threonine-proline sites in mitosis. A combination of site-directed 
      mutagenesis in a tagged raptor cDNA and analysis with a series of new 
      phospho-specific antibodies generated against different sites in raptor revealed 
      that Serine 696 and Threonine 706 represent two key sites in raptor 
      phosphorylated in mitosis. We demonstrate that the mitotic cyclin-dependent 
      kinase cdc2/CDK1 is the kinase responsible for phosphorylating these sites, and 
      its mitotic partner Cyclin B efficiently coimmunoprecipitates with raptor in 
      mitotic cells. CONCLUSIONS/SIGNIFICANCE: This study demonstrates that the key 
      mTOR binding partner raptor is directly phosphorylated during mitosis by cdc2. 
      This reinforces previous studies suggesting that mTOR activity is highly 
      regulated and important for mitotic progression, and points to a direct 
      modulation of the mTORC1 complex during mitosis.
FAU - Gwinn, Dana M
AU  - Gwinn DM
AD  - Molecular and Cell Biology Laboratory, Dulbecco Center for Cancer Research, La 
      Jolla, California, United States of America.
FAU - Asara, John M
AU  - Asara JM
FAU - Shaw, Reuben J
AU  - Shaw RJ
LA  - eng
GR  - P01 CA120964/CA/NCI NIH HHS/United States
GR  - R01 DK080425/DK/NIDDK NIH HHS/United States
GR  - HHMI/Howard Hughes Medical Institute/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
PT  - Research Support, Non-U.S. Gov't
DEP - 20100212
PL  - United States
TA  - PLoS One
JT  - PloS one
JID - 101285081
RN  - 0 (Adaptor Proteins, Signal Transducing)
RN  - 0 (CRTC2 protein, human)
RN  - 0 (Cyclin B)
RN  - 0 (RPTOR protein, human)
RN  - 0 (Regulatory-Associated Protein of mTOR)
RN  - 0 (Transcription Factors)
RN  - 2ZD004190S (Threonine)
RN  - 452VLY9402 (Serine)
RN  - EC 2.7.11.22 (CDC2 Protein Kinase)
RN  - EC 2.7.11.22 (CDK1 protein, human)
RN  - EC 2.7.11.22 (Cyclin-Dependent Kinases)
SB  - IM
MH  - Adaptor Proteins, Signal Transducing/genetics/*metabolism
MH  - Amino Acid Sequence
MH  - Binding Sites/genetics
MH  - CDC2 Protein Kinase/genetics/*metabolism
MH  - Cell Line
MH  - Cell Line, Tumor
MH  - Cyclin B/genetics/metabolism
MH  - Cyclin-Dependent Kinases
MH  - HeLa Cells
MH  - Humans
MH  - Immunoblotting
MH  - Immunoprecipitation
MH  - Mass Spectrometry
MH  - *Mitosis
MH  - Molecular Sequence Data
MH  - Mutagenesis, Site-Directed
MH  - Phosphorylation
MH  - Protein Binding
MH  - Regulatory-Associated Protein of mTOR
MH  - Sequence Homology, Amino Acid
MH  - Serine/metabolism
MH  - Threonine/metabolism
MH  - Transcription Factors/metabolism
MH  - Transfection
PMC - PMC2820552
COIS- Competing Interests: The authors have declared that no competing interests exist.
EDAT- 2010/02/20 06:00
MHDA- 2010/10/01 06:00
PMCR- 2010/02/12
CRDT- 2010/02/20 06:00
PHST- 2009/12/17 00:00 [received]
PHST- 2010/01/08 00:00 [accepted]
PHST- 2010/02/20 06:00 [entrez]
PHST- 2010/02/20 06:00 [pubmed]
PHST- 2010/10/01 06:00 [medline]
PHST- 2010/02/12 00:00 [pmc-release]
AID - 09-PONE-RA-14993 [pii]
AID - 10.1371/journal.pone.0009197 [doi]
PST - epublish
SO  - PLoS One. 2010 Feb 12;5(2):e9197. doi: 10.1371/journal.pone.0009197.