Abstract
SIN3 corepressor complexes play important roles in both normal development and breast cancer. Mammalian cells have two paralogs of SIN3 (SIN3A and SIN3B) that are encoded by distinct genes and have unique functions in many developmental processes. However, specific roles for SIN3A and SIN3B in breast cancer progression have not been characterized. We generated stable knockdown cells of SIN3 paralogs individually and in combination using three non-overlapping shRNA. Stable knockdown of SIN3B caused a significant decrease in transwell invasion through Matrigel and decreased the number of invasive colonies when grown in a 3D extracellular matrix. Conversely, stable knockdown of SIN3A significantly increased transwell invasion and increased the number of invasive colonies. These results were corroborated in vivo in which SIN3B knockdown significantly decreased and SIN3A knockdown increased experimental lung metastases. RNA sequencing was used to identify unique targets and biological pathways that were altered upon knockdown of SIN3A compared to SIN3B. Additionally, we analyzed microarray data sets to identify correlations of SIN3A and SIN3B expression with survival in patients with breast cancer. These data sets indicated that high mRNA expression of SIN3A as well as low mRNA expression of SIN3B correlates with longer relapse free survival specifically in patients with triple negative breast cancer which corresponds with our in vitro and in vivo data. These results demonstrate key functional differences between SIN3 paralogs in regulating the process of breast cancer metastasis and suggest metastasis suppressive roles of SIN3A and metastasis promoting roles of SIN3B.
Highlights
The stage-specific five-year relative survival rate for patients with distant breast cancer metastases at diagnosis is approximately 25%; a rate that has not significantly changed in the past two decades and is compared to a relative five-year survival rate of near 100% for patients with localized disease [1]
To test whether SIN3 expression is required for breast cancer metastasis, we generated cell lines with stable knockdown of the SIN3 paralogs individually and in combination
We quantified the number of invasive versus non-invasive colonies compared to the vector control in MDA-MB-231 cells: SIN3A knockdown resulted in a 38% increase and SIN3B knockdown resulted in a 49% decrease of invasive colonies (Figure 2F)
Summary
The stage-specific five-year relative survival rate for patients with distant breast cancer metastases at diagnosis is approximately 25%; a rate that has not significantly changed in the past two decades and is compared to a relative five-year survival rate of near 100% for patients with localized disease [1]. It has become increasingly clear that many of these metastasis-associated genes are regulated epigenetically by chromatin remodeling complexes that play important roles in normal development [4,5,6]. A key epigenetic regulator that is required for normal development and has been implicated in breast cancer progression is SIN3 (Switch-Independent 3) [7, 8]. SIN3 is a scaffolding protein that regulates gene transcription through chromatin modification by recruiting histone deacetylases (HDACs) to function at particular sites of the genome, typically leading to transcriptional silencing several genes have been shown to be activated [9,10,11]. Mammalian cells have two SIN3 paralogs (SIN3A and SIN3B) that are encoded by genes on chromosomes 15 and 19 respectively. The proteins are 52% identical and 68% similar with regard to amino acid sequence and mouse Sin3A is more similar to human www.impactjournals.com/oncotarget
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