Cell Signaling Technology

Product Pathways - Vesicle Trafficking

Eps15 (D3K8R) Rabbit mAb #12460

Applications Reactivity Sensitivity MW (kDa) Isotype
W IP IF-IC H M R Mk (C) (X) (Dg) (Hr) Endogenous 138 Rabbit IgG

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IF-IC=Immunofluorescence (Immunocytochemistry)
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  C=Chicken  X=Xenopus  Dg=Dog  Hr=Horse
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Eps15 (D3K8R) Rabbit mAb recognizes endogenous levels of total Eps15 protein. Based upon sequence alignment, this antibody is predicted to react with both Eps15a and Eps15b. This antibody does not cross-react with Eps15R.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro630 of human Eps15 protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Eps15 (D3K8R) Rabbit mAb.

IP

IP

Immunoprecipitation of Eps15 from MCF7 cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or Eps15 (D3K8R) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot was performed using Eps15 (D3K8R) Rabbit mAb.

IF-IC

IF-IC

Confocal immunofluorescent analysis of A-431 cells, serum-starved (left) or treated with Human Epidermal Growth Factor (hEGF) #8916 (100 ng/mL, 1 hr, 4ºC; right), using Eps15 (D3K8R) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).


Background

Eps15 (EGFR pathway substrate 15) was originally discovered as a substrate for the kinase activity of EGFR (1). Eps15 has a tripartite structure comprising an amino terminal portion, which contains three evolutionarily conserved EH protein-protein interaction domains, a central putative coiled-coil region required for constitutive oligmerization, and a carboxy terminal domain containing multiple copies of the amino acid triplet Asp-Pro-Phe that constitute the AP2 binding domain. The carboxy terminal domain also contains two ubiquitin interaction motifs (UIMs), the last of which is indespensible for Eps15 binding to ubiquitin (1). Several lines of evidence support a role for Eps15 in clathrin-mediated endocytosis, including the endocytosis of synaptic vesicles. Eps15 binds to AP2 as well as other proteins involved in endocytosis and/or synaptic vesicle recycling, such as synaptojanin1 and epsin. Furthermore, Eps15 colocalizes with markers of the plasma membrane clathrin-coated pits and vesicles (2). Eps15 regulates the endosomal trafficking of c-Met (3) and EGFR (4), possibly by recruiting the ubiquitinated receptors to the rims of clathrin-coated pits through interaction between the ubiquitin tag and its UIMs.The EPS15 gene yields two isoforms that are believed to reside in distinct subcellular locations and are thus implicated in different facets of endosomal trafficking (5). Human EPS15 has been mapped to chromosome 1p31-p32, a region displaying several nonrandom chromosomal abnormalities, including deletions in neuroblastoma and translocations in acute lymphoblastic and myeloid leukemias. Research has shown two translocations t(1;11)(p32;q11) are found in rare cases of myeloid leukemia where the Eps15 gene was fused to the HRX gene, resulting in two reciprocal fusion genes (6).

  1. Fazioli, F. et al. (1993) Mol Cell Biol 13, 5814-28.
  2. Tebar, F. et al. (1996) J Biol Chem 271, 28727-30.
  3. Parachoniak, C.A. and Park, M. (2009) J Biol Chem 284, 8382-94.
  4. Torrisi, M.R. et al. (1999) Mol Biol Cell 10, 417-34.
  5. Roxrud, I. et al. (2008) J Cell Biol 180, 1205-18.
  6. Bernard, O.A. et al. (1994) Oncogene 9, 1039-45.

Application References

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For Research Use Only. Not For Use In Diagnostic Procedures.

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