Limits...
Intracellular CD24 disrupts the ARF-NPM interaction and enables mutational and viral oncogene-mediated p53 inactivation.

Wang L, Liu R, Ye P, Wong C, Chen GY, Zhou P, Sakabe K, Zheng X, Wu W, Zhang P, Jiang T, Bassetti MF, Jube S, Sun Y, Zhang Y, Zheng P, Liu Y - Nat Commun (2015)

Bottom Line: CD24 competitively inhibits ARF binding to NPM, resulting in decreased ARF, increase MDM2 and decrease levels of p53 and the p53 target p21/CDKN1A.CD24 silencing prevents functional inactivation of p53 by both somatic mutation and viral oncogenes, including the SV40 large T antigen and human papilloma virus 16 E6-antigen.In support of the functional interaction between CD24 and p53, in silico analyses reveal that TP53 mutates at a higher rate among glioma and prostate cancer samples with higher CD24 mRNA levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA.

ABSTRACT
CD24 is overexpressed in nearly 70% human cancers, whereas TP53 is the most frequently mutated tumour-suppressor gene that functions in a context-dependent manner. Here we show that both targeted mutation and short hairpin RNA (shRNA) silencing of CD24 retard the growth, progression and metastasis of prostate cancer. CD24 competitively inhibits ARF binding to NPM, resulting in decreased ARF, increase MDM2 and decrease levels of p53 and the p53 target p21/CDKN1A. CD24 silencing prevents functional inactivation of p53 by both somatic mutation and viral oncogenes, including the SV40 large T antigen and human papilloma virus 16 E6-antigen. In support of the functional interaction between CD24 and p53, in silico analyses reveal that TP53 mutates at a higher rate among glioma and prostate cancer samples with higher CD24 mRNA levels. These data provide a general mechanism for functional inactivation of ARF and reveal an important cellular context for genetic and viral inactivation of TP53.

Show MeSH

Related in: MedlinePlus

CD24 promotes growth of DU145 cells.(a) CD24 mRNA levels in three human prostate cancer cell lines are displaced as % of glyceraldehyde 3-phosphate dehydrogenase (GAPDH). (b) Efficient gene silencing of the CD24 gene based on the protein (insert, representative images that have been reproduced three times) and RNA levels. (c) ShRNA silencing eliminates cell surface CD24 expression as determined by flow cytometry. Data shown are representative profiles and have been reproduced three times. (d) ShRNA silencing of CD24 reduces the growth rate as measured by the 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide assay. Means and s.d. of the relative cell proliferation are shown on the given days. The study has been reproduced three times. (e) Reduced growth rate of tumour cells based on colony size. Data shown are representative images and have been reproduced three times. (f) ShRNA silencing of CD24 reduced the tumour growth in the Il2rg−/−NOD.SCID (NSG) mice. The mice received a subcutaneous injection of 5 × 106 tumour cells. Data shown are means and s.d. of the tumour volumes. (g) Weights of tumours at day 39. Error bars, s.d. Scr n=14, Sh1, n=9, Sh2 n=9 for f and g. Substantially similar data were obtained when the cells were injected into nu/nu mice.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4300525&req=5

f3: CD24 promotes growth of DU145 cells.(a) CD24 mRNA levels in three human prostate cancer cell lines are displaced as % of glyceraldehyde 3-phosphate dehydrogenase (GAPDH). (b) Efficient gene silencing of the CD24 gene based on the protein (insert, representative images that have been reproduced three times) and RNA levels. (c) ShRNA silencing eliminates cell surface CD24 expression as determined by flow cytometry. Data shown are representative profiles and have been reproduced three times. (d) ShRNA silencing of CD24 reduces the growth rate as measured by the 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide assay. Means and s.d. of the relative cell proliferation are shown on the given days. The study has been reproduced three times. (e) Reduced growth rate of tumour cells based on colony size. Data shown are representative images and have been reproduced three times. (f) ShRNA silencing of CD24 reduced the tumour growth in the Il2rg−/−NOD.SCID (NSG) mice. The mice received a subcutaneous injection of 5 × 106 tumour cells. Data shown are means and s.d. of the tumour volumes. (g) Weights of tumours at day 39. Error bars, s.d. Scr n=14, Sh1, n=9, Sh2 n=9 for f and g. Substantially similar data were obtained when the cells were injected into nu/nu mice.

Mentions: As the first step to assess the function of CD24 in the growth of human prostate cancer cell lines, we analysed the expression of CD24 mRNA and protein from three cell lines, DU145, LNCaP and PC3. As shown in Fig. 3a, DU145 expressed the highest levels of CD24 mRNA, whereas LNCaP did not express detectable levels. A moderate level of CD24 mRNA was observed in the PC3 cell line. To determine the function of CD24 in DU145 proliferation, we used two independent shRNAs to silence CD24 expression. As shown in Fig. 3b,c, both shRNAs reduced CD24 mRNA levels by approximately fourfold and CD24 protein levels to barely detectable levels. In both 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide (Fig. 3d) and colony formation (Fig. 3e) assays, a significant reduction in DU145 growth was observed after silencing CD24. Consistent with this observation, the growth rates of the DU145 tumour in immunodeficient Il2Rg−/−Nod.Scid (NSG) mice (Fig. 3f) and nu/nu mice (Supplementary Fig. 5) were substantially reduced (Fig. 3f). The role for CD24 in tumour growth is further substantiated by a significant reduction in terminal tumour weight (Fig. 3g).


Intracellular CD24 disrupts the ARF-NPM interaction and enables mutational and viral oncogene-mediated p53 inactivation.

Wang L, Liu R, Ye P, Wong C, Chen GY, Zhou P, Sakabe K, Zheng X, Wu W, Zhang P, Jiang T, Bassetti MF, Jube S, Sun Y, Zhang Y, Zheng P, Liu Y - Nat Commun (2015)

CD24 promotes growth of DU145 cells.(a) CD24 mRNA levels in three human prostate cancer cell lines are displaced as % of glyceraldehyde 3-phosphate dehydrogenase (GAPDH). (b) Efficient gene silencing of the CD24 gene based on the protein (insert, representative images that have been reproduced three times) and RNA levels. (c) ShRNA silencing eliminates cell surface CD24 expression as determined by flow cytometry. Data shown are representative profiles and have been reproduced three times. (d) ShRNA silencing of CD24 reduces the growth rate as measured by the 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide assay. Means and s.d. of the relative cell proliferation are shown on the given days. The study has been reproduced three times. (e) Reduced growth rate of tumour cells based on colony size. Data shown are representative images and have been reproduced three times. (f) ShRNA silencing of CD24 reduced the tumour growth in the Il2rg−/−NOD.SCID (NSG) mice. The mice received a subcutaneous injection of 5 × 106 tumour cells. Data shown are means and s.d. of the tumour volumes. (g) Weights of tumours at day 39. Error bars, s.d. Scr n=14, Sh1, n=9, Sh2 n=9 for f and g. Substantially similar data were obtained when the cells were injected into nu/nu mice.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4300525&req=5

f3: CD24 promotes growth of DU145 cells.(a) CD24 mRNA levels in three human prostate cancer cell lines are displaced as % of glyceraldehyde 3-phosphate dehydrogenase (GAPDH). (b) Efficient gene silencing of the CD24 gene based on the protein (insert, representative images that have been reproduced three times) and RNA levels. (c) ShRNA silencing eliminates cell surface CD24 expression as determined by flow cytometry. Data shown are representative profiles and have been reproduced three times. (d) ShRNA silencing of CD24 reduces the growth rate as measured by the 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide assay. Means and s.d. of the relative cell proliferation are shown on the given days. The study has been reproduced three times. (e) Reduced growth rate of tumour cells based on colony size. Data shown are representative images and have been reproduced three times. (f) ShRNA silencing of CD24 reduced the tumour growth in the Il2rg−/−NOD.SCID (NSG) mice. The mice received a subcutaneous injection of 5 × 106 tumour cells. Data shown are means and s.d. of the tumour volumes. (g) Weights of tumours at day 39. Error bars, s.d. Scr n=14, Sh1, n=9, Sh2 n=9 for f and g. Substantially similar data were obtained when the cells were injected into nu/nu mice.
Mentions: As the first step to assess the function of CD24 in the growth of human prostate cancer cell lines, we analysed the expression of CD24 mRNA and protein from three cell lines, DU145, LNCaP and PC3. As shown in Fig. 3a, DU145 expressed the highest levels of CD24 mRNA, whereas LNCaP did not express detectable levels. A moderate level of CD24 mRNA was observed in the PC3 cell line. To determine the function of CD24 in DU145 proliferation, we used two independent shRNAs to silence CD24 expression. As shown in Fig. 3b,c, both shRNAs reduced CD24 mRNA levels by approximately fourfold and CD24 protein levels to barely detectable levels. In both 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide (Fig. 3d) and colony formation (Fig. 3e) assays, a significant reduction in DU145 growth was observed after silencing CD24. Consistent with this observation, the growth rates of the DU145 tumour in immunodeficient Il2Rg−/−Nod.Scid (NSG) mice (Fig. 3f) and nu/nu mice (Supplementary Fig. 5) were substantially reduced (Fig. 3f). The role for CD24 in tumour growth is further substantiated by a significant reduction in terminal tumour weight (Fig. 3g).

Bottom Line: CD24 competitively inhibits ARF binding to NPM, resulting in decreased ARF, increase MDM2 and decrease levels of p53 and the p53 target p21/CDKN1A.CD24 silencing prevents functional inactivation of p53 by both somatic mutation and viral oncogenes, including the SV40 large T antigen and human papilloma virus 16 E6-antigen.In support of the functional interaction between CD24 and p53, in silico analyses reveal that TP53 mutates at a higher rate among glioma and prostate cancer samples with higher CD24 mRNA levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA.

ABSTRACT
CD24 is overexpressed in nearly 70% human cancers, whereas TP53 is the most frequently mutated tumour-suppressor gene that functions in a context-dependent manner. Here we show that both targeted mutation and short hairpin RNA (shRNA) silencing of CD24 retard the growth, progression and metastasis of prostate cancer. CD24 competitively inhibits ARF binding to NPM, resulting in decreased ARF, increase MDM2 and decrease levels of p53 and the p53 target p21/CDKN1A. CD24 silencing prevents functional inactivation of p53 by both somatic mutation and viral oncogenes, including the SV40 large T antigen and human papilloma virus 16 E6-antigen. In support of the functional interaction between CD24 and p53, in silico analyses reveal that TP53 mutates at a higher rate among glioma and prostate cancer samples with higher CD24 mRNA levels. These data provide a general mechanism for functional inactivation of ARF and reveal an important cellular context for genetic and viral inactivation of TP53.

Show MeSH
Related in: MedlinePlus