PMID- 10652316
OWN - NLM
STAT- MEDLINE
DCOM- 20000302
LR  - 20190508
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 275
IP  - 5
DP  - 2000 Feb 4
TI  - Mapping of Eps15 domains involved in its targeting to clathrin-coated pits.
PG  - 3288-95
AB  - Clathrin-coated pit (CCP) formation occurs as a result of the targeting and
      assembly of cytosolic coat proteins, mainly the plasma membrane
      clathrin-associated protein complex (AP-2) and clathrin, to the intracellular
      face of the plasma membrane. In the present study, the mechanisms by which Eps15,
      an AP-2-binding protein, is targeted to CCPs was analyzed by following the
      intracellular localization of Eps15 mutants fused to the green fluorescent
      protein. Our previous results indicated that the N-terminal Eps15 homology (EH)
      domains are required for CCP targeting. We now show that EH domains are, however,
      not sufficient for targeting to CCPs. Similarly, neither the central coiled-coil 
      nor the C-terminal AP-2 binding domains were able to address green fluorescent
      protein to CCPs. Thus, targeting of Eps15 to CCPs likely results from the
      collaboration between EH domains and another domain of the protein. An Eps15
      mutant lacking the coiled-coil domain localized to CCPs showing that Eps15
      dimerization is not strictly required. In contrast, Eps15 mutants lacking all
      AP-2 binding sites showed a dramatic decrease in plasma membrane staining,
      showing that AP-2 binding sites, together with EH domains, play an important role
      in targeting Eps15 into CCPs. Finally, the effect of the Eps15 mutants on
      clathrin-dependent endocytosis was tested by both immunofluorescence and flow
      cytometry. The results obtained showed that inhibition of transferrin uptake was 
      observed only with mutants able to interfere with CCP assembly.
FAU - Benmerah, A
AU  - Benmerah A
AD  - INSERM E9925, Faculte Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris, 
      Cedex 15, France. benmerah@necker.fr
FAU - Poupon, V
AU  - Poupon V
FAU - Cerf-Bensussan, N
AU  - Cerf-Bensussan N
FAU - Dautry-Varsat, A
AU  - Dautry-Varsat A
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  - 0 (Adaptor Proteins, Signal Transducing)
RN  - 0 (Calcium-Binding Proteins)
RN  - 0 (EPS15 protein, human)
RN  - 0 (Intracellular Signaling Peptides and Proteins)
RN  - 0 (Luminescent Proteins)
RN  - 0 (Phosphoproteins)
RN  - 147336-22-9 (Green Fluorescent Proteins)
SB  - IM
MH  - Adaptor Proteins, Signal Transducing
MH  - Calcium-Binding Proteins/chemistry/genetics/*metabolism
MH  - Coated Pits, Cell-Membrane/*metabolism
MH  - Endocytosis
MH  - Green Fluorescent Proteins
MH  - HeLa Cells
MH  - Humans
MH  - Intracellular Signaling Peptides and Proteins
MH  - Luminescent Proteins
MH  - Mutation
MH  - Phosphoproteins/chemistry/genetics/*metabolism
MH  - Signal Transduction
EDAT- 2000/02/01 09:00
MHDA- 2000/03/04 09:00
CRDT- 2000/02/01 09:00
PHST- 2000/02/01 09:00 [pubmed]
PHST- 2000/03/04 09:00 [medline]
PHST- 2000/02/01 09:00 [entrez]
AID - 10.1074/jbc.275.5.3288 [doi]
PST - ppublish
SO  - J Biol Chem. 2000 Feb 4;275(5):3288-95. doi: 10.1074/jbc.275.5.3288.