SUPT5H Post-Transcriptional Silencing Modulates PIN1 Expression, Inhibits Tumorigenicity, and Induces Apoptosis of Human Breast Cancer Cells.

BACKGROUND/AIMS: Breast cancer (BrCa) is one of the most common cancers and a highly heterogenous disease, both at the pathological and molecular levels. A common element for the progression of cancer is the presence of aberrant transcription. Targeting the misregulation of transcription may serve as a tool for cancer therapeutics. SUPT5H (Suppressor of Ty 5 homolog) is a highly conserved RNA polymerase II-associated transcription elongation and processivity factor. However, few studies have examined the relationship between SUPT5H and cancer.

METHODS

Yeast two-hybrid and colocalization by immunofluorescence were performed to investigate protein-protein interaction. Colony formation assay, CTG assay, and crystal violet assays were performed for cell viability, clonogenicity, and cell proliferation study. Data mining was performed for expression analysis of SUPT5H in breast cancers. Flow cytometry was performed for the assessment of cell cycle and apoptosis. The Transwell chambers were employed for the migration and invasion assays. Quantitative real-time polymerase chain reaction (qRT PCR) and Western blotting were performed to measure the mRNA and protein levels of SUPT5H and other markers related to viability, migration, cell cycle, and apoptosis. Silent mutations were generated for rescue experiments. The biological function of SUPT5H was investigated through siRNA depletion of SUPT5H mRNA in vitro.

RESULTS

We showed that SUPT5H is upregulated in breast cancer tissue as compared with the adjacent normal tissue in breast cancer patients. In human breast cancer cells, the levels of SUPT5H and PIN1 are positively correlated with each other. Our biochemical analysis showed that PIN1 interacts with SUPT5H through WW domain, that was required to promote SUPT5H protein stability. Depletion of SUPT5H by siRNA technology reduced the tumorigenic and metastatic properties, promoted s-phase cell cycle arrest and apoptosis of MDA-MB-231 cells. Moreover, depletion of SUPT5H abrogated MAPK molecules thereby regulates the oncogenic behavior of breast cancer cells.

CONCLUSION

Our findings demonstrated an essential role of SUPT5H in BrCa tumorigenicity by regulating the expression levels of genes that control proliferation, migration, cell cycle, and apoptosis of breast cancer MDA-MB-231 cells.