A monoclonal antibody against p53 cross-reacts with processing bodies

Authors
<div class="autor_fcen" id="8534">Thomas, M.G.</div>; <div class="autor_fcen" id="5196">Luchelli, L.</div>; Pascual, M.; Gottifredi, V.; <div class="autor_fcen" id="982">Boccaccio, G.L.</div>
Publication Year
2012
Language
English
Format
article
Status
Published version
Description
The p53 tumor suppressor protein is an important regulator of cell proliferation and apoptosis. p53 can be found in the nucleus and in the cytosol, and the subcellular location is key to control p53 function. In this work, we found that a widely used monoclonal antibody against p53, termed Pab 1801 (Pan antibody 1801) yields a remarkable punctate signal in the cytoplasm of several cell lines of human origin. Surprisingly, these puncta were also observed in two independent p53-null cell lines. Moreover, the foci stained with the Pab 1801 were present in rat cells, although Pab 1801 recognizes an epitope that is not conserved in rodent p53. In contrast, the Pab 1801 nuclear staining corresponded to genuine p53, as it was upregulated by p53-stimulating drugs and absent in p53-null cells. We identified the Pab 1801 cytoplasmic puncta as P Bodies (PBs), which are involved in mRNA regulation. We found that, in several cell lines, including U2OS, WI38, SK-N-SH and HCT116, the Pab 1801 puncta strictly colocalize with PBs identified with specific antibodies against the PB components Hedls, Dcp1a, Xrn1 or Rck/p54. PBs are highly dynamic and accordingly, the Pab 1801 puncta vanished when PBs dissolved upon treatment with cycloheximide, a drug that causes polysome stabilization and PB disruption. In addition, the knockdown of specific PB components that affect PB integrity simultaneously caused PB dissolution and the disappearance of the Pab 1801 puncta. Our results reveal a strong cross-reactivity of the Pab 1801 with unknown PB component(s). This was observed upon distinct immunostaining protocols, thus meaning a major limitation on the use of this antibody for p53 imaging in the cytoplasm of most cell types of human or rodent origin. © 2012 Thomas et al.
Fil:Thomas, M.G. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.
Fil:Luchelli, L. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.
Fil:Boccaccio, G.L. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.
Source
PLoS ONE 2012;7(5)
Subject
cell marker
cycloheximide
decapping enzyme 1a
decapping enzyme 1b
decapping enzyme 2
epitope
exoribonuclease
exoribonuclease 1
messenger RNA
monoclonal antibody
pantropic antibody 1801
protein 4ET
protein Hedls
protein p53
protein p54
small interfering RNA
unclassified drug
epitope
protein p53
TP53 protein, human
animal cell
animal tissue
article
cell component
cell disruption
cell line
cell stimulation
cell strain HCT116
cellular distribution
concentration (parameters)
controlled study
cross reaction
dissolution
Drosophila
embryo
fetus
gene control
gene silencing
genetic transfection
human
human cell
image analysis
immunohistochemistry
intracellular signaling
nonhuman
polysome
processing body
protein analysis
protein localization
rat
upregulation
animal
antibody specificity
chemistry
cytoplasm
immunology
metabolism
Sprague Dawley rat
tumor cell line
Rattus
Rodentia
Animals
Antibodies, Monoclonal, Murine-Derived
Antibody Specificity
Cell Line, Tumor
Cytoplasm
Epitopes
Humans
Immunohistochemistry
Rats
Rats, Sprague-Dawley
Tumor Suppressor Protein p53
Access level
Open access
License
http://creativecommons.org/licenses/by/2.5/ar
Repository
Biblioteca Digital (UBA-FCEN)
Institution
Universidad Nacional de Buenos Aires. Facultad de Ciencias Exactas y Naturales
OAI Identifier
snrd:HASHd669aec9837f3e3dabf480