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AurkA controls self-renewal of breast cancer-initiating cells promoting wnt3a stabilization through suppression of miR-128.

Eterno V, Zambelli A, Villani L, Tuscano A, Manera S, Spitaleri A, Pavesi L, Amato A - Sci Rep (2016)

Bottom Line: In our study, AurkA knock-down reduced wnt3a mRNA and suppressed metastatic signature of MDA-MB-231 cells.As a consequence, the amount of BCICs and their migratory capability dramatically decreased.In addition, it suggests a new therapeutic strategy taking advantage of miR-128 to suppress AurkA-Wnt3a signaling.

View Article: PubMed Central - PubMed

Affiliation: Lab of Experimental Oncology &Pharmacogenomics IRCCS Fondazione "Salvatore Maugeri", Pavia, Italy.

ABSTRACT
AurkA overexpression was previously found in breast cancer and associated to its ability in controlling chromosome segregation during mitosis, however whether it may affect breast cancer cells, endorsed with stem properties (BCICs), is still unclear. Surprisingly, a strong correlation between AurkA expression and β-catenin localization in breast cancer tissues suggested a link between AurkA and Wnt signaling. In our study, AurkA knock-down reduced wnt3a mRNA and suppressed metastatic signature of MDA-MB-231 cells. As a consequence, the amount of BCICs and their migratory capability dramatically decreased. Conversely, wnt3a mRNA stabilization and increased CD44(+)/CD24(low/-) subpopulation was found in AurkA-overexpressing MCF7 cells. In vivo, AurkA-overexpressing primary breast cancer cells showed higher tumorigenic properties. Interestingly, we found that AurkA suppressed the expression of miR-128, inhibitor of wnt3a mRNA stabilization. Namely, miR-128 suppression realized after AurkA binding to Snail. Remarkably, a strong correlation between AurkA and miR-128 expression in breast cancer tissues confirmed our findings. This study provides novel insights into an undisclosed role for the kinase AurkA in self-renewal and migration of BCICs affecting response to cancer therapies, metastatic spread and recurrence. In addition, it suggests a new therapeutic strategy taking advantage of miR-128 to suppress AurkA-Wnt3a signaling.

No MeSH data available.


Related in: MedlinePlus

Snail mediates repression of miR-128 in response to Aurka overexpression.(A) AurkA overexpression does not promote Snail protein increase as found in MCF-AurkA (AurkA+) versus control cells (empty) Evaluation of Snail protein levels in MCF-7 and MCF-AurkA. In contrast, AurkA knock-down severely impaired Snail protein levels in MDA-shAurkA (sh5, sh7, sh8) and KBr2-shAurkA (shAurkA) in comparison with control cells (empty). (B) AurkA antibody pulls down Snail in MCF7 and MDA-MB-231 cells after a co-immunoprecipitation assay (IP anti-AurkA). As control, protein lysates from both cell lines were tested for Snail protein levels (control). A marker was loaded in the last lane as a control of molecular weight. (C) AurkA overexpression in MCF-7 cells induced nuclear translocation of Snail (MCF, AurkA+). Control cells show a moderate cytoplasmic staining (MCF, empty). Nuclear accumulation of Snail in MDA-MB-231 cells (MDA, empty) was inhibited by AurkA knock-down (MDA, shAurkA). Nuclei were counterstained with DAPI. Collectively our findings supports a signaling pathway showing that AurkA may promote nuclear translocation of Snail to repress miR-128 gene transcription. As results, wnt3a mRNA accumulates increasing Wnt3a protein levels. (D) On the left, Q-PCR confirmed association between AurkA and miR-128 levels in 32 human tissues from breast cancer patients. Asterisk highlights samples were association is missing. On the right, Graphic representation of of samples (percentage, %) showing a strong correlation AurkA-mir-128 (AurkA > 1.5/miR-128 < 1.5 and AurkA < 1.5/miR-128 > 1.5), or where AurkA-miR-128 correlation is missing (AurkA > 1.5/miR-128 > 1.5 and AurkA < 1.5/miR-128 < 1.5).
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f6: Snail mediates repression of miR-128 in response to Aurka overexpression.(A) AurkA overexpression does not promote Snail protein increase as found in MCF-AurkA (AurkA+) versus control cells (empty) Evaluation of Snail protein levels in MCF-7 and MCF-AurkA. In contrast, AurkA knock-down severely impaired Snail protein levels in MDA-shAurkA (sh5, sh7, sh8) and KBr2-shAurkA (shAurkA) in comparison with control cells (empty). (B) AurkA antibody pulls down Snail in MCF7 and MDA-MB-231 cells after a co-immunoprecipitation assay (IP anti-AurkA). As control, protein lysates from both cell lines were tested for Snail protein levels (control). A marker was loaded in the last lane as a control of molecular weight. (C) AurkA overexpression in MCF-7 cells induced nuclear translocation of Snail (MCF, AurkA+). Control cells show a moderate cytoplasmic staining (MCF, empty). Nuclear accumulation of Snail in MDA-MB-231 cells (MDA, empty) was inhibited by AurkA knock-down (MDA, shAurkA). Nuclei were counterstained with DAPI. Collectively our findings supports a signaling pathway showing that AurkA may promote nuclear translocation of Snail to repress miR-128 gene transcription. As results, wnt3a mRNA accumulates increasing Wnt3a protein levels. (D) On the left, Q-PCR confirmed association between AurkA and miR-128 levels in 32 human tissues from breast cancer patients. Asterisk highlights samples were association is missing. On the right, Graphic representation of of samples (percentage, %) showing a strong correlation AurkA-mir-128 (AurkA > 1.5/miR-128 < 1.5 and AurkA < 1.5/miR-128 > 1.5), or where AurkA-miR-128 correlation is missing (AurkA > 1.5/miR-128 > 1.5 and AurkA < 1.5/miR-128 < 1.5).

Mentions: Indeed, we found that MDA-shAurkA and KBr2-shAurkA showed decreased snail in comparison with control cells (Fig. 6A), surprisingly no significant change was found in MCF-AurkA+ cells which showed Snail protein levels similar to control cells (Fig. 6A).


AurkA controls self-renewal of breast cancer-initiating cells promoting wnt3a stabilization through suppression of miR-128.

Eterno V, Zambelli A, Villani L, Tuscano A, Manera S, Spitaleri A, Pavesi L, Amato A - Sci Rep (2016)

Snail mediates repression of miR-128 in response to Aurka overexpression.(A) AurkA overexpression does not promote Snail protein increase as found in MCF-AurkA (AurkA+) versus control cells (empty) Evaluation of Snail protein levels in MCF-7 and MCF-AurkA. In contrast, AurkA knock-down severely impaired Snail protein levels in MDA-shAurkA (sh5, sh7, sh8) and KBr2-shAurkA (shAurkA) in comparison with control cells (empty). (B) AurkA antibody pulls down Snail in MCF7 and MDA-MB-231 cells after a co-immunoprecipitation assay (IP anti-AurkA). As control, protein lysates from both cell lines were tested for Snail protein levels (control). A marker was loaded in the last lane as a control of molecular weight. (C) AurkA overexpression in MCF-7 cells induced nuclear translocation of Snail (MCF, AurkA+). Control cells show a moderate cytoplasmic staining (MCF, empty). Nuclear accumulation of Snail in MDA-MB-231 cells (MDA, empty) was inhibited by AurkA knock-down (MDA, shAurkA). Nuclei were counterstained with DAPI. Collectively our findings supports a signaling pathway showing that AurkA may promote nuclear translocation of Snail to repress miR-128 gene transcription. As results, wnt3a mRNA accumulates increasing Wnt3a protein levels. (D) On the left, Q-PCR confirmed association between AurkA and miR-128 levels in 32 human tissues from breast cancer patients. Asterisk highlights samples were association is missing. On the right, Graphic representation of of samples (percentage, %) showing a strong correlation AurkA-mir-128 (AurkA > 1.5/miR-128 < 1.5 and AurkA < 1.5/miR-128 > 1.5), or where AurkA-miR-128 correlation is missing (AurkA > 1.5/miR-128 > 1.5 and AurkA < 1.5/miR-128 < 1.5).
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Related In: Results  -  Collection

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Show All Figures
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f6: Snail mediates repression of miR-128 in response to Aurka overexpression.(A) AurkA overexpression does not promote Snail protein increase as found in MCF-AurkA (AurkA+) versus control cells (empty) Evaluation of Snail protein levels in MCF-7 and MCF-AurkA. In contrast, AurkA knock-down severely impaired Snail protein levels in MDA-shAurkA (sh5, sh7, sh8) and KBr2-shAurkA (shAurkA) in comparison with control cells (empty). (B) AurkA antibody pulls down Snail in MCF7 and MDA-MB-231 cells after a co-immunoprecipitation assay (IP anti-AurkA). As control, protein lysates from both cell lines were tested for Snail protein levels (control). A marker was loaded in the last lane as a control of molecular weight. (C) AurkA overexpression in MCF-7 cells induced nuclear translocation of Snail (MCF, AurkA+). Control cells show a moderate cytoplasmic staining (MCF, empty). Nuclear accumulation of Snail in MDA-MB-231 cells (MDA, empty) was inhibited by AurkA knock-down (MDA, shAurkA). Nuclei were counterstained with DAPI. Collectively our findings supports a signaling pathway showing that AurkA may promote nuclear translocation of Snail to repress miR-128 gene transcription. As results, wnt3a mRNA accumulates increasing Wnt3a protein levels. (D) On the left, Q-PCR confirmed association between AurkA and miR-128 levels in 32 human tissues from breast cancer patients. Asterisk highlights samples were association is missing. On the right, Graphic representation of of samples (percentage, %) showing a strong correlation AurkA-mir-128 (AurkA > 1.5/miR-128 < 1.5 and AurkA < 1.5/miR-128 > 1.5), or where AurkA-miR-128 correlation is missing (AurkA > 1.5/miR-128 > 1.5 and AurkA < 1.5/miR-128 < 1.5).
Mentions: Indeed, we found that MDA-shAurkA and KBr2-shAurkA showed decreased snail in comparison with control cells (Fig. 6A), surprisingly no significant change was found in MCF-AurkA+ cells which showed Snail protein levels similar to control cells (Fig. 6A).

Bottom Line: In our study, AurkA knock-down reduced wnt3a mRNA and suppressed metastatic signature of MDA-MB-231 cells.As a consequence, the amount of BCICs and their migratory capability dramatically decreased.In addition, it suggests a new therapeutic strategy taking advantage of miR-128 to suppress AurkA-Wnt3a signaling.

View Article: PubMed Central - PubMed

Affiliation: Lab of Experimental Oncology &Pharmacogenomics IRCCS Fondazione "Salvatore Maugeri", Pavia, Italy.

ABSTRACT
AurkA overexpression was previously found in breast cancer and associated to its ability in controlling chromosome segregation during mitosis, however whether it may affect breast cancer cells, endorsed with stem properties (BCICs), is still unclear. Surprisingly, a strong correlation between AurkA expression and β-catenin localization in breast cancer tissues suggested a link between AurkA and Wnt signaling. In our study, AurkA knock-down reduced wnt3a mRNA and suppressed metastatic signature of MDA-MB-231 cells. As a consequence, the amount of BCICs and their migratory capability dramatically decreased. Conversely, wnt3a mRNA stabilization and increased CD44(+)/CD24(low/-) subpopulation was found in AurkA-overexpressing MCF7 cells. In vivo, AurkA-overexpressing primary breast cancer cells showed higher tumorigenic properties. Interestingly, we found that AurkA suppressed the expression of miR-128, inhibitor of wnt3a mRNA stabilization. Namely, miR-128 suppression realized after AurkA binding to Snail. Remarkably, a strong correlation between AurkA and miR-128 expression in breast cancer tissues confirmed our findings. This study provides novel insights into an undisclosed role for the kinase AurkA in self-renewal and migration of BCICs affecting response to cancer therapies, metastatic spread and recurrence. In addition, it suggests a new therapeutic strategy taking advantage of miR-128 to suppress AurkA-Wnt3a signaling.

No MeSH data available.


Related in: MedlinePlus