Abstract
Abstract
Recent findings have shown that the Heat Shock Protein 90 (Hsp90) co-chaperone UNC45A is overexpressed in ovarian and breast cancers. Previously, we have shown that UNC45A is a centrosomal protein essential for cervical tumor cell growth through activation of the checkpoint kinase 1 (ChK1). In this report, we further examined the role of UNC45A in breast tumorigenesis using a variety of biochemical and cell biology techniques and animal models. We confirmed that UNC45A is highly overexpressed in human breast-infiltrating ductal carcinomas as compared to adjacent normal tissues. Silencing UNC45A in vitro blocked the proliferation of all breast cancer subtypes and drastically reduced tumor growth of the triple negative MDA-MB-231 cell line implanted in mammary fat pads of NOD/SCID mice. However, loss of UNC45A did not affect the proliferation of normal mammary cells. Remarkably, UNC45A becomes more nuclear in human cancer tissues and cancer cell lines as compared to normal tissues and non-transformed Hs578Bst and HME mammary cell lines, respectively. This suggests an important nuclear function for UNC45A during tumorigenesis. Microarray analysis of mRNA from Hs578T cells showed that loss of UNC45A alters the expression of 121 genes, involved in cancer and cellular development and growth networks. Relevant to cell proliferation, we found that Nek7 gene was significantly repressed upon silencing UNC45A, which was validated by RTqPCR and Western blot analyses in multiple breast cancer cell lines. Nek7 is a member of the NIMA (never in mitosis, gene A) family of serine/threonine kinases. It plays a key role in centrosomal separation during mitosis. This correlates neatly with our observation that loss of UNC45A causes a centrosomal separation defect, cell proliferation arrest and death of breast cancer cell lines. ChIP experiments showed that UNC45A binds to the promoter of the Nek7 gene, suggesting direct transcriptional regulation. Interestingly, the UNC45A sequence contains four LxxLL motifs, which are thought to be signatures for co-activator binding to nuclear receptors. Furthermore, computational analysis identified two glucocorticoid response elements (GRE) consensus sequences in the Nek7 promoter, suggesting its transcriptional regulation by the glucocorticoid receptor (GR). This hypothesis was further strengthened by a significant decrease in the mRNA and protein levels of Nek7 upon silencing GR. Thus, our data suggest that UNC45A functions as a GR co-activator to control Nek7 gene transcription. Consistent with this, immunoprecipitation experiments confirmed that UNC45A and GR form endogenous complexes, and treatment of Hs578T and MCF7 cell lines with dexamethasone upregulates Nek7 mRNA and protein levels. In conclusion our data strongly support the premise that UNC45A promotes Nek7 transcription through activation of GR, and thus controls centrosomal separation and cancer cell proliferation.
Citation Format: Yasmeen Jilani, Nada H. Eisa, Kashish Kainth, Sumin Lu, Nehal M. Elsherbiny, Laila A. Eissa, Mamdouh M. Elshishtawy, Hasan Korkaya, Abdeljabar El Andaloussi, Ahmed Chadli. The co-chaperone UNC45A controls cancer cell proliferation through Nek7 and centrosomal separation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4493. doi:10.1158/1538-7445.AM2017-4493