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Diverse effects of ANXA7 and p53 on LNCaP prostate cancer cells are associated with regulation of SGK1 transcription and phosphorylation of the SGK1 target FOXO3A.

Srivastava M, Leighton X, Starr J, Eidelman O, Pollard HB - Biomed Res Int (2014)

Bottom Line: In addition, p53 reduced the expression of aberrant SGK1 protein form in LNCaP.Using Ingenuity Pathway Analysis and p53-signature genes, we elucidated the role of distinct SGK1/FOXO3A-associated regulation in p53 versus ANXA7 responses and proposed that aberrant SGK1 could affect reciprocal SGK1-FOXO3A-Akt regulation.Thus, the failure of the cell growth regulator p53 versus the phospholipid-binding ANXA7 could be potentially attributed to its diverse effects on SGK1-FOXO3A-Akt pathway in the PTEN-deficient LNCaP.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Physiology and Genetics and Institute for Molecular Medicine, Uniformed Services University of Health Sciences (USUHS), School of Medicine, Bethesda, MD 20814, USA.

ABSTRACT
Tumor suppressor function of the calcium/phospholipid-binding Annexin-A7 (ANXA7) has been shown in Anxa7-deficient mice and validated in human cancers. In the androgen-resistant prostate cancer cells, ANXA7 and p53 showed similar cytotoxicity levels. However, in the androgen-sensitive LNCaP, ANXA7 greatly exceeded the p53-induced cytotoxicity. We hypothesized that the p53 underperformance in LNCaP could be due to the involvement of p53-responsive SGK1 and FOXO3A. In this study, we show that p53 failed to match programmed cell death (PCD) and G1-arrest that were induced by ANXA7 in LNCaP. WT-ANXA7 preserved total FOXO3A expression with no hyperphosphorylation that could enable FOXO3A nuclear translocation and proapoptotic transcription. In contrast, in the p53-transfected LNCaP cells with maintained cell proliferation, the phosphorylated (but not total) FOXO3A fraction was increased implying a predominantly cytoplasmic localization and, subsequently, a lack of FOXO3A proapoptotic transcription. In addition, p53 reduced the expression of aberrant SGK1 protein form in LNCaP. Using Ingenuity Pathway Analysis and p53-signature genes, we elucidated the role of distinct SGK1/FOXO3A-associated regulation in p53 versus ANXA7 responses and proposed that aberrant SGK1 could affect reciprocal SGK1-FOXO3A-Akt regulation. Thus, the failure of the cell growth regulator p53 versus the phospholipid-binding ANXA7 could be potentially attributed to its diverse effects on SGK1-FOXO3A-Akt pathway in the PTEN-deficient LNCaP.

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LNCaP cells transfected with vector, WT/DN-ANXA7, or p53. Protein synexpression was assessed by Western blotting. Arrows designate FOXO3A-S318/321 hyperphosphorylation in the p53-transfected LNCaP.
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fig3: LNCaP cells transfected with vector, WT/DN-ANXA7, or p53. Protein synexpression was assessed by Western blotting. Arrows designate FOXO3A-S318/321 hyperphosphorylation in the p53-transfected LNCaP.

Mentions: While similar tumor suppressor effects of WT-ANXA7 and p53 in DU145 were accompanied by similar RB-E2F profiles, the lack of RB1 dephosphorylation and E2F induction by p53 compared to WT-ANXA7 contributed to the p53 insufficiency in androgen-sensitive LNCaP [4]. In the PTEN-mutant LNCaP with a constitutively active Akt which is expected to phosphorylate FOXO3A [5], p53 activation can further hyperphosphorylate FOXO3A [6], thus preventing the proapoptotic FOXO3A transcription. Hence, we compared WT/DN-ANXA7 and p53 effects on the expression of FOXO3A in LNCaP. As shown in Figure 3, FOXO3A phosphorylation in LNCaP was distinctly affected by WT/DN-ANXA7 and p53 at the Ser318/321 site. Consistent with the lack of tumor suppressor effects, p53 drastically hyperphosphorylated FOXO3A-Ser318/321 in LNCaP, but not in other prostate cells (data not shown). In addition, p53 and DN- (but not WT-) ANXA7 enhanced total FOXO3A degradation: 85 kDa was reduced and 20 kDa increased. In the meantime, the expression of full-length SGK1 was not changed, while LMW SGK1 products (including aberrant <45 kDa) were enhanced by WT-ANXA7 and reduced by p53. These results suggest that the LMW-SGK1 enhancement by ANXA7 in LNCaP was accompanied by the reduced FOXO3A-S318/321 phosphorylation which indicated a nuclear translocation of FOXO3A. In contrast, the p53-induced FOXO3A-S318/321 hyperphosphorylation indicated the enhancement of FOXO3A cytoplasmic retention and deactivation in LNCaP.


Diverse effects of ANXA7 and p53 on LNCaP prostate cancer cells are associated with regulation of SGK1 transcription and phosphorylation of the SGK1 target FOXO3A.

Srivastava M, Leighton X, Starr J, Eidelman O, Pollard HB - Biomed Res Int (2014)

LNCaP cells transfected with vector, WT/DN-ANXA7, or p53. Protein synexpression was assessed by Western blotting. Arrows designate FOXO3A-S318/321 hyperphosphorylation in the p53-transfected LNCaP.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4016907&req=5

fig3: LNCaP cells transfected with vector, WT/DN-ANXA7, or p53. Protein synexpression was assessed by Western blotting. Arrows designate FOXO3A-S318/321 hyperphosphorylation in the p53-transfected LNCaP.
Mentions: While similar tumor suppressor effects of WT-ANXA7 and p53 in DU145 were accompanied by similar RB-E2F profiles, the lack of RB1 dephosphorylation and E2F induction by p53 compared to WT-ANXA7 contributed to the p53 insufficiency in androgen-sensitive LNCaP [4]. In the PTEN-mutant LNCaP with a constitutively active Akt which is expected to phosphorylate FOXO3A [5], p53 activation can further hyperphosphorylate FOXO3A [6], thus preventing the proapoptotic FOXO3A transcription. Hence, we compared WT/DN-ANXA7 and p53 effects on the expression of FOXO3A in LNCaP. As shown in Figure 3, FOXO3A phosphorylation in LNCaP was distinctly affected by WT/DN-ANXA7 and p53 at the Ser318/321 site. Consistent with the lack of tumor suppressor effects, p53 drastically hyperphosphorylated FOXO3A-Ser318/321 in LNCaP, but not in other prostate cells (data not shown). In addition, p53 and DN- (but not WT-) ANXA7 enhanced total FOXO3A degradation: 85 kDa was reduced and 20 kDa increased. In the meantime, the expression of full-length SGK1 was not changed, while LMW SGK1 products (including aberrant <45 kDa) were enhanced by WT-ANXA7 and reduced by p53. These results suggest that the LMW-SGK1 enhancement by ANXA7 in LNCaP was accompanied by the reduced FOXO3A-S318/321 phosphorylation which indicated a nuclear translocation of FOXO3A. In contrast, the p53-induced FOXO3A-S318/321 hyperphosphorylation indicated the enhancement of FOXO3A cytoplasmic retention and deactivation in LNCaP.

Bottom Line: In addition, p53 reduced the expression of aberrant SGK1 protein form in LNCaP.Using Ingenuity Pathway Analysis and p53-signature genes, we elucidated the role of distinct SGK1/FOXO3A-associated regulation in p53 versus ANXA7 responses and proposed that aberrant SGK1 could affect reciprocal SGK1-FOXO3A-Akt regulation.Thus, the failure of the cell growth regulator p53 versus the phospholipid-binding ANXA7 could be potentially attributed to its diverse effects on SGK1-FOXO3A-Akt pathway in the PTEN-deficient LNCaP.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Physiology and Genetics and Institute for Molecular Medicine, Uniformed Services University of Health Sciences (USUHS), School of Medicine, Bethesda, MD 20814, USA.

ABSTRACT
Tumor suppressor function of the calcium/phospholipid-binding Annexin-A7 (ANXA7) has been shown in Anxa7-deficient mice and validated in human cancers. In the androgen-resistant prostate cancer cells, ANXA7 and p53 showed similar cytotoxicity levels. However, in the androgen-sensitive LNCaP, ANXA7 greatly exceeded the p53-induced cytotoxicity. We hypothesized that the p53 underperformance in LNCaP could be due to the involvement of p53-responsive SGK1 and FOXO3A. In this study, we show that p53 failed to match programmed cell death (PCD) and G1-arrest that were induced by ANXA7 in LNCaP. WT-ANXA7 preserved total FOXO3A expression with no hyperphosphorylation that could enable FOXO3A nuclear translocation and proapoptotic transcription. In contrast, in the p53-transfected LNCaP cells with maintained cell proliferation, the phosphorylated (but not total) FOXO3A fraction was increased implying a predominantly cytoplasmic localization and, subsequently, a lack of FOXO3A proapoptotic transcription. In addition, p53 reduced the expression of aberrant SGK1 protein form in LNCaP. Using Ingenuity Pathway Analysis and p53-signature genes, we elucidated the role of distinct SGK1/FOXO3A-associated regulation in p53 versus ANXA7 responses and proposed that aberrant SGK1 could affect reciprocal SGK1-FOXO3A-Akt regulation. Thus, the failure of the cell growth regulator p53 versus the phospholipid-binding ANXA7 could be potentially attributed to its diverse effects on SGK1-FOXO3A-Akt pathway in the PTEN-deficient LNCaP.

Show MeSH
Related in: MedlinePlus