Abstract
The p160/Steroid Receptor Coactivators SRC-1, SRC-2/GRIP1, and SRC-3/AIB1 are important regulators of Estrogen Receptor alpha (ERα) activity. However, whereas the functions of SRC-1 and SRC-3 in breast tumourigenesis have been extensively studied, little is known about the role of SRC-2. Previously, we reported that activation of the cAMP-dependent protein kinase, PKA, facilitates ubiquitination and proteasomal degradation of SRC-2 which in turn leads to inhibition of SRC-2-coactivation of ERα and changed expression of the ERα target gene, pS2. Here we have characterized the global program of transcription in SRC-2-depleted MCF-7 breast cancer cells using short-hairpin RNA technology, and in MCF-7 cells exposed to PKA activating agents. In order to identify genes that may be regulated through PKA-induced downregulation of SRC-2, overlapping transcriptional targets in response to the respective treatments were characterized. Interestingly, we observed decreased expression of several breast cancer tumour suppressor genes (e.g., TAGLN, EGR1, BCL11b, CAV1) in response to both SRC-2 knockdown and PKA activation, whereas the expression of a number of other genes implicated in cancer progression (e.g., RET, BCAS1, TFF3, CXCR4, ADM) was increased. In line with this, knockdown of SRC-2 also stimulated proliferation of MCF-7 cells. Together, these results suggest that SRC-2 may have an antiproliferative function in breast cancer cells.
Highlights
The Steroid Receptor Coactivator (SRC) family, known as p160 proteins, consist of the three members, SRC-1 (NCOA1) [1], SRC-2 (NCOA2/GRIP1/TIF2) [2,3], and SRC-3 (NCOA3/ AIB1/ACTR/RAC-3/pCIP/TRAM-1) [4,5,6,7,8]
We have shown that activation of protein kinase (PKA) leads to increased ubiquitination and subsequent degradation of SRC-2, which in turn leads to inhibition of Estrogen Receptor alpha (ERa) transactivation function [35,36]
Since our results indicated that KD of SRC-2 changes the expression of estrogen-responsive genes known to be involved in carcinogenesis, we wanted to examine whether KD of SRC-2 affected the real time growth of MCF-7 cells by using the xCELLigence System
Summary
The Steroid Receptor Coactivator (SRC) family, known as p160 proteins, consist of the three members, SRC-1 (NCOA1) [1], SRC-2 (NCOA2/GRIP1/TIF2) [2,3], and SRC-3 (NCOA3/ AIB1/ACTR/RAC-3/pCIP/TRAM-1) [4,5,6,7,8]. Even though depletion of SRC-2 in MCF-7 cells has been shown to decrease estrogen-dependent ERa transactivation function, loss of SRC-2 does not seem to affect estrogen-dependent proliferation of MCF-7 cells [29] This is in contrast to studies of SRC-3-depleted MCF-7 cells in which the estrogen-dependent proliferation of the cells was significantly reduced [29,30,31,32]. Distinct PTMs of the SRC members play a crucial role in controlling their intracellular levels and functions, which may have significant impact on breast carcinogenesis and response to endocrine treatment in breast cancer patients [18]. In previous studies we have demonstrated that activation of the cAMP-dependent protein kinase (PKA) signalling pathway targets SRC-2 coactivator function through its ubiquitination and proteasomal degradation.
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