PMID- 10037681
OWN - NLM
STAT- MEDLINE
DCOM- 19990330
LR  - 20210209
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 274
IP  - 10
DP  - 1999 Mar 5
TI  - Interaction of the small G protein RhoA with the C terminus of human
      phospholipase D1.
PG  - 6035-8
AB  - Mammalian phosphatidylcholine-specific phospholipase D1 (PLD1) is a signal
      transduction-activated enzyme thought to function in multiple cell biological
      settings including the regulation of membrane vesicular trafficking. PLD1 is
      activated by the small G proteins, ADP-ribosylation factor (ARF) and RhoA, and by
      protein kinase C-alpha (PKC-alpha). This stimulation has been proposed to involve
      direct interaction and to take place at a distinct site in PLD1 for each
      activator. In the present study, we employed the yeast two-hybrid system to
      attempt to identify these sites. Successful interaction of ARF and PKC-alpha with
      PLD1 was not achieved, but a C-terminal fragment of human PLD1 (denoted "D4")
      interacted with the active mutant of RhoA, RhoAVal-14. Deletion of the CAAX box
      from RhoAVal-14 decreased the strength of the interaction, suggesting that lipid 
      modification of RhoA is important for efficient binding to PLD1. The specificity 
      of the interaction was validated by showing that the PLD1 D4 fragment interacts
      with glutathione S-transferase-RhoA in vitro in a GTP-dependent manner and that
      it associates with RhoAVal-14 in COS-7 cells, whereas the N-terminal two-thirds
      of PLD1 does not. Finally, we show that recombinant D4 peptide inhibits
      RhoA-stimulated PLD1 activation but not ARF- or PKC-alpha-stimulated PLD1
      activation. These results conclusively demonstrate that the C-terminal region of 
      PLD1 contains the RhoA-binding site and suggest that the ARF and PKC interactions
      occur elsewhere in the protein.
FAU - Yamazaki, M
AU  - Yamazaki M
AD  - Department of Life Science, Tokyo Institute of Technology, Yokohama 226-8501,
      Japan.
FAU - Zhang, Y
AU  - Zhang Y
FAU - Watanabe, H
AU  - Watanabe H
FAU - Yokozeki, T
AU  - Yokozeki T
FAU - Ohno, S
AU  - Ohno S
FAU - Kaibuchi, K
AU  - Kaibuchi K
FAU - Shibata, H
AU  - Shibata H
FAU - Mukai, H
AU  - Mukai H
FAU - Ono, Y
AU  - Ono Y
FAU - Frohman, M A
AU  - Frohman MA
FAU - Kanaho, Y
AU  - Kanaho Y
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - EC 3.1.4.4 (Phospholipase D)
RN  - EC 3.1.4.4 (phospholipase D1)
RN  - EC 3.6.1.- (GTP-Binding Proteins)
RN  - EC 3.6.5.2 (rhoA GTP-Binding Protein)
SB  - IM
MH  - Binding Sites
MH  - GTP-Binding Proteins/*chemistry/metabolism
MH  - Humans
MH  - Phospholipase D/*chemistry/metabolism
MH  - Protein Binding
MH  - rhoA GTP-Binding Protein
EDAT- 1999/02/26 00:00
MHDA- 1999/02/26 00:01
CRDT- 1999/02/26 00:00
PHST- 1999/02/26 00:00 [pubmed]
PHST- 1999/02/26 00:01 [medline]
PHST- 1999/02/26 00:00 [entrez]
AID - 10.1074/jbc.274.10.6035 [doi]
AID - S0021-9258(19)87550-6 [pii]
PST - ppublish
SO  - J Biol Chem. 1999 Mar 5;274(10):6035-8. doi: 10.1074/jbc.274.10.6035.