p21Waf1 expression is regulated by nuclear intermediate filament vimentin in neuroblastoma.
BACKGROUND
Human neuroblastoma (NB) cell lines may present with either one of the so-called S-and N-subtypes. We have previously reported a strong correlation between protein expression levels of vimentin, an S-subtype marker, and the p21Waf1 cyclin-dependent kinase inhibitor. We here investigated whether this correlation extend to the mRNA level in NB cell lines as well as in patients' tumors. We also further explored the relationship between expression of vimentin and p21, by asking whether vimentin could regulate p21 expression.


METHODS
Vimentin and p21 mRNA levels in NB cell lines as well as in patients' tumors (n = 77) were quantified using Q-PCR. Q-PCR data obtained from tumors of high risk NB patients (n = 40) were analyzed in relation with the overall survival using the Log-rank Kaplan-Meier estimation. siRNA-mediated depletion or overexpression of vimentin in highly or low expressing vimentin cell lines, respectively, followed by protein expression and promoter activation assays were used to assess the role of vimentin in modulating p21 expression.


RESULTS
We extend the significant correlation between vimentin and p21 expression to the mRNA level in NB cell lines as well as in patients' tumors. Overall survival analysis from Q-PCR data obtained from tumors of high risk patients suggests that lower levels of p21 expression could be associated with a poorer outcome. Our data additionally indicate that the correlation observed between p21 and vimentin expression levels results from p21 transcriptional activity being regulated by vimentin. Indeed, downregulating vimentin resulted in a significant decrease in p21 mRNA and protein expression as well as in p21 promoter activity. Conversely, overexpressing vimentin triggered an increase in p21 promoter activity in cells with a nuclear expression of vimentin.


CONCLUSIONS
Our results suggest that p21 mRNA tumor expression level could represent a refined prognostic factor for high risk NB patients. Our data also show that vimentin regulates p21 transcription; this is the first demonstration of a gene regulating function for this type III-intermediate filament.
Mergui
Xénia
X
Université Paris Sud-11, CNRS, UMR 8126, Institut de Cancérologie Gustave Roussy, 114 rue Edouard Vaillant, Villejuif F-94805, France. benard@igr.fr
Puiffe
Marie-Line
ML
Valteau-Couanet
Dominique
D
Lipinski
Marc
M
Bénard
Jean
J
Amor-Guéret
Mounira
M
D000293:Adolescent; D002467:Cell Nucleus / Q000378:metabolism* / Q000473:pathology; D002648:Child; D002675:Child, Preschool; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008297:Male; D009447:Neuroblastoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D011379:Prognosis; D011401:Promoter Regions, Genetic / Q000235:genetics; D034741:RNA, Small Interfering / Q000494:pharmacology; D015996:Survival Rate; D014407:Tumor Cells, Cultured; D014746:Vimentin / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D034741:RNA, Small Interfering; D014746:Vimentin
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