Abstract
Here, we report a novel non-epigenetic function of histone deacetylase (HDAC) 8 in activating cancer stem cell (CSC)-like properties in breast cancer cells by enhancing the stability of Notch1 protein. The pan-HDAC inhibitors AR-42 and SAHA, and the class I HDAC inhibitor depsipeptide, suppressed mammosphere formation and other CSC markers by reducing Notch1 expression in MDA-MB-231 and SUM-159 cells. Interrogation of individual class I isoforms (HDAC1-3 and 8) using si/shRNA-mediated knockdown, ectopic expression and/or pharmacological inhibition revealed HDAC8 to be the primary mediator of this drug effect. This suppression of Notch1 in response to HDAC8 inhibition was abrogated by the proteasome inhibitor MG132 and siRNA-induced silencing of Fbwx7, indicating Notch1 suppression occurred through proteasomal degradation. However, co-immunoprecipitation analysis indicated that HDAC8 did not form complexes with Notch1 and HDAC inhibition had no effect on Notch1 acetylation. In a xenograft tumor model, the tumorigenicity of breast cancer cells was decreased by HDAC8 knockdown. These findings suggest the therapeutic potential of HDAC8 inhibition to suppress Notch1 signaling in breast cancer.
Highlights
We report a novel non-epigenetic function of HDAC8 in regulating cancer stem cell (CSC)-like properties in breast cancer cells by maintaining the stability of Notch1
D protein, which might underlie the reported ability of histone deacetylase (HDAC) inhibitors to activate Notch1 signaling in cancer cells
A tumorigenic role for HDAC8 has been suggested by its high expression levels in childhood neuroblastoma
Summary
Knockdown of HDAC8 in MDA-MB-231 cells, using two different shRNAs (#71 and #74) that displayed no cross-inhibition of the other three class I HDAC isoforms, led to concomitant decreases in the expression of Notch and the CSC markers CD133, CD44 and Kruppel-like factor 4 (KLF4) (Figure 2B). This HDAC8 knockdown-mediated inhibition of Notch signaling, as shown by reduced expression of Notch 1 and its downstream targets NICD, Nestin, and BMI-1, decreased the abilities of MDA-MB-231 and SUM159 cells to form mammospheres as compared to control cells (Figure 3A). With the exception of BT474 and MDA-MB-468 cells, HDAC8 expression was positively correlated with that of Notch, with SUM159 cells exhibiting the highest abundance of both HDAC8 and Notch, followed by T47D and MDA-MB-231
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