Steroid Receptor RNA Activator Protein Research Articles

The steroid receptor RNA activator protein (SRAP) controls cancer cell migration/motility

The steroid receptor RNA activator gene (SRA1) produces both a functional RNA (SRA) and a protein (SRAP), whose exact physiological roles remain unknown. To identify cellular processes regulated by SRAP we compared the transcriptome of Hela and MDA-MB-231 cancer cells upon depletion of the SRA/SRAP transcripts or overexpression of the SRAP protein. RNA-seq and Ontology analyses pinpointed cellular movement as potentially regulated by SRAP.Using live cell imaging, we found that SRA/SRAP depletion and SRAP overexpression lead respectively to a decrease and increase in cancer cell motility.Our results highlight for the first time a link existing between SRA1 gene expression and cell motility.

Increased steroid receptor RNA activator protein (SRAP) accompanied by decreased estrogen receptor-beta (ER-β) levels during the malignant transformation of endometriosis associated ovarian clear cell carcinoma

The modulating attributes of steroid receptor RNA activator protein (SRAP) on steroid receptors have been shown in some types of tumor cells. There is compelling evidence to suggest that this molecule may play a critical role in the development of the tumor. However, little has been reported on its expression in endometriosis associated ovarian clear cell carcinoma (EAOCCC). In order to investigate the role of SRAP and estrogen receptors (ERs) in EAOCCC, we have analyzed the distribution of these proteins in the malignant transformation tissues and endometrioma tissues by immunohistochemistry. Our results revealed that the positive ratio of ER-β expression was gradually reduced during the malignant transformation from endometriosis to atypical endometriosis to clear cell carcinoma. Conversely, during the process, a gradual increase in SRAP expression was observed. Furthermore, there is a negative relationship between the expressions of these two molecules. Overall an increase in SRAP and a reduction in ER-β expression might be associated with malignant transformation of EAOCCC.

Influence of ovarian endometrioma on expression of steroid receptor RNA activator, estrogen receptors, vascular endothelial growth factor, and thrombospondin 1 in the surrounding ovarian tissues.

This study investigates the influence of ovarian endometrioma on expression of steroid receptor RNA activator (SRA), estrogen receptors (ERs), vascular endothelial growth factor (VEGF), and thrombospondin 1 (TSP-1) in the surrounding ovarian tissues. Taken from the women with ovarian endometrioma and mature teratoma during laparoscopy, the biopsies were analyzed by real-time polymerase chain reaction and Western blot. Our results indicated that ovarian tissues surrounding endometrioma had lower SRA and ER-α levels but higher SRA protein (SRAP) and ER-β levels than ovarian endometrioma. With lower VEGF levels and higher TSP-1 levels, the surrounding ovarian tissues showed higher expression levels of SRA, SRAP, ER-α, and ER-β in the ovarian endometrioma group when compared to the controls. These data showed that ovarian endometrioma increases SRA, ERs, and TSP-1 but decreases VEGF levels in the surrounding ovarian tissues, suggesting that abnormal expression of these molecules may affect biological behaviors of ovarian endometrioma.

Structure and Function of Steroid Receptor RNA Activator Protein, the Proposed Partner of SRA Noncoding RNA

In a widely accepted model, the steroid receptor RNA activator protein (SRA protein; SRAP) modulates the transcriptional regulatory activity of SRA RNA by binding a specific stem–loop of SRA. We first confirmed that SRAP is present in the nucleus as well as the cytoplasm of MCF-7 breast cancer cells, where it is expressed at the level of about 105 molecules per cell. However, our SRAP–RNA binding experiments, both in vitro with recombinant protein and in cultured cells with plasmid-expressed protein and RNA, did not reveal a specific interaction between SRAP and SRA. We determined the crystal structure of the carboxy-terminal domain of human SRAP and found that it does not have the postulated RRM (RNA recognition motif). The structure is a five-helix bundle that is distinct from known RNA-binding motifs and instead is similar to the carboxy-terminal domain of the yeast spliceosome protein PRP18, which stabilizes specific protein–protein interactions within a multisubunit mRNA splicing complex. SRA binding experiments with this domain gave negative results. Transcriptional regulation by SRA/SRAP was examined with siRNA knockdown. Effects on both specific estrogen-responsive genes and genes identified by RNA-seq as candidates for regulation were examined in MCF-7 cells. Only a small effect (~20% change) on one gene resulting from depletion of SRA/SRAP could be confirmed. We conclude that the current model for SRAP function must be reevaluated; we suggest that SRAP may function in a different context to stabilize specific intermolecular interactions in the nucleus.

Steroid receptor RNA activator protein (SRAP) expression as a prognostic factor in ER+ human breast tumors

The steroid receptor RNA activator protein (SRAP) is a newly described protein modulating the activity of multiple transcription factors including the estrogen receptor (ER). We have recently reported the immunodetection by Western blot of multiple SRAP peptides in breast tissue. High expression of these peptides, assessed by tissue micro-array (TMA) analysis, was associated with poor prognosis in patients whose primary tumors were ER positive (ER+). In such studies, it is recognized that intensity as well as specificity of the signal detected directly depends upon the antibody used as well as the position of the epitope recognized. To confirm the potential relevance of SRAP as a new prognostic factor, it is critical to establish whether similar results are obtained with independent antibodies. Two commercial anti-SRAP antibodies (742A and 743A), respectively, recognizing the N- and C-terminal extremity of the protein, were first used to analyze by Western blot SRAP expression in protein extracts from frozen breast tumor tissue sections. These antibodies were further used to investigate by immunohistochemistry (IHC) SRAP location in paraffin-embedded breast tumors. Comparative TMA analysis of 170 ER+ tumors was eventually performed in order to establish the potential associations existing between SRAP expression and clinical outcome. Multiple SRAP peptides were differentially detected by Western blot. Both antibodies led to similar nuclear and cytoplasmic staining in breast tissue section. A solid correlation was found (Spearman r = 0.46, P < 0.001) between 742A and 743A IHC scores. Results from both antibodies independently showed that dividing expression levels into lower 25 percentile, 26-75 percentile, and highest 25 percentile demonstrated a hazard ratio (HR) of 1.82 (P = 0.0042) for 742A antibody and 1.35 (P = 0.14) for 743A antibody. When both scores are combined, double high expressor (by 742A and 743A) was associated with a poor prognosis of breast-cancer-specific survival (Mantel-Cox: P = 0.005, HR = 2.24). Overall, our data suggest the existence in breast tumor tissue of multiple SRAP-like peptides. Assessing their expression in primary breast tumors can predict clinical outcome in ER+ breast cancer patients.

Expression of both Estrogen Receptor-beta 1 (ER-β1) and its co-regulator Steroid Receptor RNA Activator Protein (SRAP) are predictive for benefit from tamoxifen therapy in patients with Estrogen Receptor-alpha (ER-α)-Negative Early Breast Cancer (EBC)

BackgroundRoles of Estrogen Receptor-beta 1 (ER-β1) and its co-regulator Steroid Receptor RNA Activator Protein (SRAP) in breast cancer remain unclear. Previously, ER-β1 and SRAP expression were found positively correlated in breast cancer and, therefore, expression of these two molecules could characterize cancers with a distinct clinical outcome. Patients and methodsER-β1 and SRAP expression was determined by immunohistochemistry (IHC) in tissue microarrays from a randomized, placebo-controlled trial (NCIC-CTG-MA12), designed to determine the benefit of tamoxifen following chemotherapy in premenopausal early breast cancer (EBC). Expression was dichotomized into low and high using median IHC scores. Relationships with survival used Cox modeling. ResultsIn the whole cohort, ER-β1 and SRAP were not prognostic. However, high ER-β1 and SRAP significantly predicted tamoxifen responsiveness [overall survival, interaction test, P = 0.03; relapse-free survival (RFS), interaction test, P = 0.01]. Stratification by ER-α-status found predictive benefit only in ER-α-negative cases. The difference in RFS between tamoxifen and placebo was greater in patients whose tumors expressed both high SRAP and ER-β1[hazard ratio = 0.07; 95% confidence interval (CI) 0.01–0.41; P = 0.003] versus those with low SRAP or ER-β1 (interaction test, P = 0.02). The interaction test was not significant in ER-α-positive cohorts. ConclusionsThis study provides evidence that both ER-β1 and SRAP could be predictive biomarkers of tamoxifen benefit in ER-α-negative premenopausal EBC.

Steroid receptor RNA activator bi-faceted genetic system: Heads or Tails?

The Steroid Receptor RNA Activator (SRA) was first identified by Lanz et al. in 1999 as a functional non-coding RNA able to co-activate steroid nuclear receptors. Since this incipient study, our understanding of SRA as a broader co-regulator of nuclear receptors as well as other transcription factors has greatly expanded. Accumulated data has now revealed the diverse roles played by this transcript in both normal biological processes such as myogenesis and adipogenesis, as well as in mechanisms underlying diseases including cardio-myopathies and cancers.Remarkably, as early as 2000, SRA isoforms were identified that were also able to code for a protein now referred to as the Steroid Receptor RNA Activator Protein (SRAP). SRA and SRAP now define a very intriguing bi-faceted genetic system, where both RNA and protein products of the same gene play specific and sometime overlapping roles in cell biology. Due to its initial molecular characterization as an RNA, most reports have in the last ten years focused on the non-coding part of this twosome. As such, only a handful of laboratories have investigated the molecular and biological roles played by SRAP. The scope of this review is to summarize and discuss our current knowledge of the molecular features and functions specifically attributable to the coding nature of the bi-faceted products of the SRA1 gene.

Steroid receptor RNA activator protein binds to and counteracts SRA RNA-mediated activation of MyoD and muscle differentiation

The steroid receptor RNA activator (SRA) has the unusual property to function as both a non-coding RNA (ncRNA) and a protein SRAP. SRA ncRNA is known to increase the activity of a range of nuclear receptors as well as the master regulator of muscle differentiation MyoD. The contribution of SRA to either a ncRNA or a protein is influenced by alternative splicing of the first intron, the retention of which disrupts the SRAP open reading frame. We reported here that the ratio between non-coding and coding SRA isoforms increased during myogenic differentiation of human satellite cells but not myotonic dystrophy patient satellite cells, in which differentiation capacity is affected. Using constructs that exclusively produce SRA ncRNA or SRAP, we demonstrated that whereas SRA ncRNA was indeed an enhancer of myogenic differentiation and myogenic conversion of non-muscle cells through the co-activation of MyoD activity, SRAP prevented this SRA RNA-dependant co-activation. Interestingly, the SRAP inhibitory effect is mediated through the interaction of SRAP with its RNA counterpart via its RRM-like domain interacting with the functional sub-structure of SRA RNA, STR7. This study thus provides a new model for SRA-mediated regulation of MyoD transcriptional activity in the promotion of normal muscle differentiation, which takes into account the nature of SRA molecules present.

The steroid receptor RNA activator protein is recruited to promoter regions and acts as a transcriptional repressor

Products of the steroid receptor RNA activator (SRA1) gene have the unusual property to function both at the RNA and the protein levels. SRA-RNA has long been known to increase the activity of multiple nuclear receptors. It has more recently been proposed than steroid receptor RNA activator protein (SRAP) also modulates steroid receptors activity. Herein, we show for the first time that SRAP physically interacts with multiple transcription factors and is recruited to specific promoter regions. Artificially recruiting SRAP to the promoter of a luciferase reporter gene under the control of the strong transcriptional activator VP16 leads to a decrease in transcription. Altogether we propose that SRAP could be a new transcriptional regulator, able to function as a repressor through direct association with promoters. Structured summaryMINT-7761068: SRAP (uniprotkb:Q9HD15) physically interacts (MI:0915) with HDAC2 (uniprotkb:Q92769) by anti bait coimmunoprecipitation (MI:0006)

Steroid Receptor RNA Activator Protein (SRAP): a potential new prognostic marker for estrogen receptor-positive/node-negative/younger breast cancer patients

IntroductionThe steroid receptor RNA activator is a functional RNA suspected to participate in the mechanisms underlying breast tumor progression. This RNA is also able to encode for a protein, Steroid Receptor RNA Activator Protein (SRAP), whose exact function remains to be determined. Our aim was to assess, in a large breast cancer cohort, whether levels of this protein could be associated with outcome or established clinical parameters.MethodsFollowing antibody validation, SRAP expression was assessed by tissue-microarray (TMA) analysis of 372 breast tumors. Clinical follow-up and parameters such as steroid receptor and node status were available for all the corresponding cases. Immunohistochemical scores were independently determined by three investigators and averaged. Statistical analyses were performed using standard univariate and multivariate tests.ResultsSRAP levels were significantly (Mann-Whitney rank sum test, P < 0.05) higher in estrogen receptor-alpha positive (ER+, n = 271), in progesterone receptor positive (PR+, n = 257) and in older patients (age > 64 years, n = 182). When considering ER+ tumors, PR+ tumors, or younger patients (≤ 64 years), cases with high SRAP expression had a significantly (Mantel-Cox test, P < 0.05) worse breast cancer specific survival (BCSS) than those with low SRAP levels. SRAP also appeared as a very powerful indicator of poor prognostic for BCSS in the subset of ER+, node negative and young breast cancer patients (Cox regression analysis, n = 60, BCSS Hazard Ratio = 8.61, P < 0.006).ConclusionsOur data suggest that SRAP levels might provide additional information on potential risk of recurrence and negative outcome in a specific set of patients with otherwise good prognosis when considering only estrogen receptor and nodal status.

Expression and function of human steroid receptor RNA activator in prostate cancer cells: role of endogenous hSRA protein in androgen receptor-mediated transcription

Steroid receptor RNA activator (SRA) was first isolated as a steroid receptor co-activator that functioned as an RNA transcript. Later, we demonstrated that SRA needs to be translated in order to co-activate androgen receptor (AR). Here, we showed that three isoforms of human SRA enhanced AR activities. Small interfering RNA against SRA suppressed AR activities in PC-3 cells transfected with pSG5AR and in LNCaP cells that have an endogenous mutated-AR. Western blot showed that SRA protein was expressed at a higher level in PC-3 than in LNCaP cells, suggesting that SRA may be related to hormone-independent growth of prostate cancer.

The steroid receptor RNA activator protein is expressed in breast tumor tissues

The steroid receptor RNA activator (SRA) was originally described as the first functional noncoding RNA able to specifically coactivate the activity of steroid receptors. We previously demonstrated the existence in breast cancer cell lines of new SRA isoforms that, as opposed to the first cloned SRA RNA, encode for a 236-amino acid protein, SRAP. To investigate the possible implications of the coding SRA RNA and SRAP expression on breast cancer progression, we examined by Western blot analysis 74 primary breast tumors of patients subsequently treated with tamoxifen. Patients whose primary tumors were positive for SRAP expression (n = 24) had a significantly (Kaplan-Meier survival curve p = 0.047) lower likelihood of dying from recurrent disease than SRAP-negative patients (n = 50). We generated 2 cell lines, SRAP-V5-High.A and SRAP-V5-High.B, by stably overexpressing SRAP in the estrogen receptor-positive MCF-7 breast cancer cell line. Transient transfection experiments, performed using a luciferase reporter gene under the control of an estrogen-responsive element, revealed decreased sensitivity to estradiol but no additional sensitivity to tamoxifen in SRAP-overexpressing cells. Overall, our data suggest that the presence of both coding SRA RNA and its corresponding SRAP modifies the activity of estrogen receptor in breast cancer cells and that SRAP could be a new clinical marker for breast cancer. Further studies are needed to define the respective mechanisms of action and the roles of SRA RNA and protein in breast tumorigenesis and tumor progression.

The steroid receptor RNA activator is the first functional RNA encoding a protein

The steroid receptor RNA activator (SRA) has previously been characterized as belonging to the growing family of functional non-coding RNAs. However, we recently reported the Western blot detection of a putative endogenous SRA protein (SRAP) in breast cancer cells. Herein, we successfully suppressed the expression of this protein through specific RNA interference assay, unequivocally confirming its existence. Moreover, using database searches and Western blot analysis, we also showed that SRAP is highly conserved among chordata. Overall, our results suggest that SRA is the first example of a new class of functional RNAs also able to encode a protein.