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Expression of C-terminal deleted p53 isoforms in neuroblastoma.

Goldschneider D, Horvilleur E, Plassa LF, Guillaud-Bataille M, Million K, Wittmer-Dupret E, Danglot G, de Thé H, Bénard J, May E, Douc-Rasy S - Nucleic Acids Res. (2006)

Bottom Line: Xirodimas, M.Saville and D.Lane (2005) Genes Dev., 19, 2122-2137].

View Article: PubMed Central - PubMed

Affiliation: Centre National de Recherche Scientifique, UMR 8126, Institut Gustave Roussy, 94805 Villejuif, France.

ABSTRACT
The tumor suppressor gene, p53, is rarely mutated in neuroblastomas (NB) at the time of diagnosis, but its dysfunction could result from a nonfunctional conformation or cytoplasmic sequestration of the wild-type p53 protein. However, p53 mutation, when it occurs, is found in NB tumors with drug resistance acquired over the course of chemotherapy. As yet, no study has been devoted to the function of the specific p53 mutants identified in NB cells. This study includes characterization and functional analysis of p53 expressed in eight cell lines: three wild-type cell lines and five cell lines harboring mutations. We identified two transcription-inactive p53 variants truncated in the C-terminus, one of which corresponded to the p53beta isoform recently identified in normal tissue by Bourdon et al. [J. C. Bourdon, K. Fernandes, F. Murray-Zmijewski, G. Liu, A. Diot, D. P. Xirodimas, M. K. Saville and D. P. Lane (2005) Genes Dev., 19, 2122-2137]. Our results show, for the first time, that the p53beta isoform is the only p53 species to be endogenously expressed in the human NB cell line SK-N-AS, suggesting that the C-terminus truncated p53 isoforms may play an important role in NB tumor development.

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Related in: MedlinePlus

p53 expression of different neuroblastoma cell lines. (A) Western blot from protein total extracts (upper panel); β-actin was used as loading control (lower panel). (B) RT–PCR using F1-R7 primers (Table 1); amplified fragment was normalized by GAPDH.
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fig1: p53 expression of different neuroblastoma cell lines. (A) Western blot from protein total extracts (upper panel); β-actin was used as loading control (lower panel). (B) RT–PCR using F1-R7 primers (Table 1); amplified fragment was normalized by GAPDH.

Mentions: We first compared the migration profiles of p53 expressed in SK-N-AS and IGR-NB8 with those expressed in the other six NB cells, SH-SY5Y, LAN-5, LAN-1, IMR-32, SK-N-BE(2) and IGR-N-91. Western blots from 50 μg of total protein extracts were revealed with p53 monoclonal antibody (DO-7). A range of profiles was identified as shown in Figure 1A. As expected, p53 extracted from the three cell lines, SH-SY5Y, LAN-5 and IMR-32, expressing wt protein (13,21) migrated at the wt position. Of particular note in these three wt p53 cell lines was an additional faint band that migrated faster than the full-length protein. The LAN-1 cells were found to be p53 deficient (9). The SK-N-BE(2) cell line showed an intense band reflecting p53 stability due to a missense mutation at codon 135 (11). As expected, due to the previously identified duplication of exons 7-8-9 (13), p53 protein migration was delayed in the IGR-N-91 cells. In contrast, the p53 protein in the SK-N-AS cell line migrated noticeably faster than the wt protein, indicating that it was smaller in size. The p53 protein in the IGR-NB8 cell line was even smaller than that in SK-N-AS.


Expression of C-terminal deleted p53 isoforms in neuroblastoma.

Goldschneider D, Horvilleur E, Plassa LF, Guillaud-Bataille M, Million K, Wittmer-Dupret E, Danglot G, de Thé H, Bénard J, May E, Douc-Rasy S - Nucleic Acids Res. (2006)

p53 expression of different neuroblastoma cell lines. (A) Western blot from protein total extracts (upper panel); β-actin was used as loading control (lower panel). (B) RT–PCR using F1-R7 primers (Table 1); amplified fragment was normalized by GAPDH.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1636465&req=5

fig1: p53 expression of different neuroblastoma cell lines. (A) Western blot from protein total extracts (upper panel); β-actin was used as loading control (lower panel). (B) RT–PCR using F1-R7 primers (Table 1); amplified fragment was normalized by GAPDH.
Mentions: We first compared the migration profiles of p53 expressed in SK-N-AS and IGR-NB8 with those expressed in the other six NB cells, SH-SY5Y, LAN-5, LAN-1, IMR-32, SK-N-BE(2) and IGR-N-91. Western blots from 50 μg of total protein extracts were revealed with p53 monoclonal antibody (DO-7). A range of profiles was identified as shown in Figure 1A. As expected, p53 extracted from the three cell lines, SH-SY5Y, LAN-5 and IMR-32, expressing wt protein (13,21) migrated at the wt position. Of particular note in these three wt p53 cell lines was an additional faint band that migrated faster than the full-length protein. The LAN-1 cells were found to be p53 deficient (9). The SK-N-BE(2) cell line showed an intense band reflecting p53 stability due to a missense mutation at codon 135 (11). As expected, due to the previously identified duplication of exons 7-8-9 (13), p53 protein migration was delayed in the IGR-N-91 cells. In contrast, the p53 protein in the SK-N-AS cell line migrated noticeably faster than the wt protein, indicating that it was smaller in size. The p53 protein in the IGR-NB8 cell line was even smaller than that in SK-N-AS.

Bottom Line: Xirodimas, M.Saville and D.Lane (2005) Genes Dev., 19, 2122-2137].

View Article: PubMed Central - PubMed

Affiliation: Centre National de Recherche Scientifique, UMR 8126, Institut Gustave Roussy, 94805 Villejuif, France.

ABSTRACT
The tumor suppressor gene, p53, is rarely mutated in neuroblastomas (NB) at the time of diagnosis, but its dysfunction could result from a nonfunctional conformation or cytoplasmic sequestration of the wild-type p53 protein. However, p53 mutation, when it occurs, is found in NB tumors with drug resistance acquired over the course of chemotherapy. As yet, no study has been devoted to the function of the specific p53 mutants identified in NB cells. This study includes characterization and functional analysis of p53 expressed in eight cell lines: three wild-type cell lines and five cell lines harboring mutations. We identified two transcription-inactive p53 variants truncated in the C-terminus, one of which corresponded to the p53beta isoform recently identified in normal tissue by Bourdon et al. [J. C. Bourdon, K. Fernandes, F. Murray-Zmijewski, G. Liu, A. Diot, D. P. Xirodimas, M. K. Saville and D. P. Lane (2005) Genes Dev., 19, 2122-2137]. Our results show, for the first time, that the p53beta isoform is the only p53 species to be endogenously expressed in the human NB cell line SK-N-AS, suggesting that the C-terminus truncated p53 isoforms may play an important role in NB tumor development.

Show MeSH
Related in: MedlinePlus