Key Spec Table
|Species Reactivity||Key Applications||Host||Format||Antibody Type|
|H, M, R||IP, WB||M||Purified||Monoclonal Antibody|
|Presentation||0.1M Tris-glycine, pH 7.4, 0.15M NaCl, 0.05% sodium azide before the addition of glycerol to 30%|
|Application||Anti-SRC-1 Antibody, clone 1135 is a Mouse Monoclonal Antibody for the detection of SRC-1 & has been validated in IP & WB.|
|Safety Information according to GHS|
|Storage and Shipping Information|
|Storage Conditions||2 years at -20°C|
|Material Size||100 µg|
|Anti-SRC-1, clone 1135||2480577|
|Anti-SRC-1, clone 1135 - 18883||18883|
|Anti-SRC-1, clone 1135 - 20690||20690|
|Anti-SRC-1, clone 1135 - 21604||21604|
|Anti-SRC-1, clone 1135 - 22759||22759|
|Anti-SRC-1, clone 1135 - 28736||28736|
|Anti-SRC-1, clone 1135 - DAM1831388||DAM1831388|
|Anti-SRC-1, clone 1135 Monoclonal Antibody||2976284|
|Reference overview||Application||Species||Pub Med ID|
|Progesterone receptor and SRC-1 participate in the regulation of VEGF, EGFR and Cyclin D1 expression in human astrocytoma cell lines.|
Olivia Tania Hernández-Hernández,Tania Karina González-García,Ignacio Camacho-Arroyo
The Journal of steroid biochemistry and molecular biology 132 2012
Astrocytomas are the most common primary brain tumors in humans. It has been reported that vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR), cyclin D1 and progesterone receptor (PR) expression levels are elevated in patients with high-grade astrocytomas. Progesterone (P) regulates astrocytomas growth through its interaction with PR, which recruits coregulatory proteins such as steroid receptor coactivator-1 (SRC-1) that are required for efficient transcriptional activation. The regulation of VEGF, EGFR and cyclin D1 expression by P in human astrocytoma cells is not known. We studied the role of PR and SRC-1 in the expression of VEGF, EGFR and cyclin D1 mediated by P in human astrocytoma cell lines grade III (U373) and IV (D54). P significantly increased VEGF and EGFR mRNA expression after 12h of treatment in D54 cells that was reflected at protein level 24h after treatment. This effect was blocked by the PR antagonist, RU 486. In U373 cells cyclin D1 mRNA and protein expression was induced by P after 6 and 8h of treatment, respectively, and this effect was blocked with RU 486. Transfection with short hairpin RNA targeting coactivator SRC-1 significantly reduced VEGF expression after 24h of treatment. Collectively, our results indicate that P regulates VEGF and EGFR expression in D54 cells and cyclin D1 expression in U373 through PR, and that SRC-1 participates in the regulation of VEGF expression.
|Estradiol increases cell growth in human astrocytoma cell lines through ERα activation and its interaction with SRC-1 and SRC-3 coactivators.|
Aliesha González-Arenas,Valeria Hansberg-Pastor,Olivia Tania Hernández-Hernández,Tania Karina González-García,Joshua Henderson-Villalpando,Diana Lemus-Hernández,Aglaé Cruz-Barrios,Mariana Rivas-Suárez,Ignacio Camacho-Arroyo
Biochimica et biophysica acta 1823 2012
Estradiol (E2) regulates several cellular functions through the interaction with estrogen receptor subtypes, ERα and ERβ, which present different functional and regulation properties. ER subtypes have been identified in human astrocytomas, the most common and aggressive primary brain tumors. We studied the role of ER subtypes in cell growth of two human astrocytoma cell lines derived from tumors of different evolution grades: U373 and D54 (grades III and IV, respectively). E2 significantly increased the number of cells in both lines and the co-administration with an ER antagonist (ICI 182, 780) significantly blocked E2 effects. ERα was the predominant subtype in both cell lines. E2 and ICI 182, 780 down-regulated ERα expression. The number of U373 and D54 cells significantly increased after PPT (ERα agonist) treatment but not after DPN (ERβ agonist) one. To determine the role of SRC-1 and SRC-3 coactivators in ERα induced cell growth, we silenced them with RNA interference. Coactivator silencing blocked the increase in cell number induced by PPT. The content of proteins involved in proliferation and metastasis was also determined after PPT treatment. Western blot analysis showed that in U373 cells the content of PR isoforms (PR-A and PR-B), EGFR, VEGF and cyclin D1 increased after PPT treatment while in D54 cells only the content of EGFR was increased. Our results demonstrate that E2 induces cell growth of human astrocytoma cell lines through ERα and its interaction with SRC-1 and SRC-3 and also suggest differential roles of ERα on cell growth depending on astrocytoma grade.
|Systems genetics analyses predict a transcription role for P2P-R: molecular confirmation that P2P-R is a transcriptional co-repressor.|
Peidis, P; Giannakouros, T; Burow, ME; Williams, RW; Scott, RE
BMC systems biology 4 14 2010
The 250 kDa P2P-R protein (also known as PACT and Rbbp6) was cloned over a decade ago and was found to bind both the p53 and Rb1 tumor suppressor proteins. In addition, P2P-R has been associated with multiple biological functions, such as mitosis, mRNA processing, translation and ubiquitination. In the current studies, the online GeneNetwork system was employed to further probe P2P-R biological functions. Molecular studies were then performed to confirm the GeneNetwork evaluations.GeneNetwork and associated gene ontology links were used to investigate the coexpression of P2P-R with distinct functional sets of genes in an adipocyte genetic reference panel of HXB/BXH recombinant strains of rats and an eye genetic reference panel of BXD recombinant inbred strains of mice. The results establish that biological networks of 75 and 135 transcription-associated gene products that include P2P-R are co-expressed in a genetically-defined manner in rat adipocytes and in the mouse eye, respectively. Of this large set of transcription-associated genes, greater than 10% are associated with hormone-mediated transcription. Since it has been previously reported that P2P-R can bind the SRC-1 transcription co-regulatory factor (steroid receptor co-activator 1, [Ncoa1]), the possible effects of P2P-R on estrogen-induced transcription were evaluated. Estrogen-induced transcription was repressed 50-70% by the transient transfection of P2P-R plasmid constructs into four different cell types. In addition, knockdown of P2P-R expression using an antisense oligonucleotide increased estrogen-mediated transcription. Co-immunoprecipitation assays confirmed that P2P-R interacts with SRC-1 and also demonstrated that P2P-R interacts with estrogen receptor alpha.The findings presented in this study provide strong support for the value of systems genetics, especially GeneNetwork, in discovering new functions of genes that can be confirmed by molecular analysis. More specifically, these data provide evidence that the expression of P2P-R co-varies in a genetically-defined manner with large transcription networks and that P2P-R can function as a co-repressor of estrogen-dependent transcription.Full Text Article
|Gonadotropin-releasing hormone-mediated phosphorylation of estrogen receptor-alpha contributes to fosB expression in mouse gonadotrophs.|
Chen, J; An, BS; Cheng, L; Hammond, GL; Leung, PC
Endocrinology 150 4583-93 2009
Estrogen receptors (ERs) are activated by their ligands as well as signaling pathways that alter ER phosphorylation in response to peptide hormones and growth factors. In pituitary gonadotrophs, GnRHs act via the type I GnRH receptor (GnRHR). Both GnRH subtypes (GnRH-I and -II) activate an estrogen response element (ERE)-driven luciferase reporter gene in LbetaT2 mouse pituitary cells, and GnRH-I is most potent in this regard. Moreover, antide (a GnRH antagonist) and a GnRHR small interfering RNA (siRNA) abrogate this effect, whereas an ERalpha antagonist (ICI 182,780) does not. The ERalpha in LbetaT2 cells is phosphorylated at Ser(118) in the nucleus and at Ser(167) in both nucleus and cytoplasm after GnRH treatments and coincided with increased ERalpha binding to its coactivator, the p300/cAMP response element-binding protein-associated factor (PCAF). Moreover, siRNA-mediated knockdown of PCAF levels attenuated GnRH-induced ERE-luciferase transactivation in these cells. Most importantly, both GnRH subtypes robustly up-regulated expression of the immediate early response gene, fosB, whereas cotreatment with ERalpha siRNA or PCAF siRNA attenuated this effect. This appears to occur at the transcriptional level because corecruitment of ERalpha and PCAF to an ERE within the endogenous fosB promoter was increased by GnRH treatments, as shown by chromatin immunoprecipitation assays. These data demonstrate that GnRH-mediated phosphorylation of ERalpha in mouse LbetaT2 pituitary cells results in its rapid association with PCAF and the transcriptional activation of fosB, and we demonstrate that this in turn likely activates other genes in pituitary cells including the FSH beta-subunit gene.
|Regulation of estrogen receptor-mediated long range transcription via evolutionarily conserved distal response elements.|
Pan, YF; Wansa, KD; Liu, MH; Zhao, B; Hong, SZ; Tan, PY; Lim, KS; Bourque, G; Liu, ET; Cheung, E
The Journal of biological chemistry 283 32977-88 2008
Nuclear signaling by estrogens rapidly induces the global recruitment of estrogen receptors (ERs) to thousands of highly specific locations in the genome. Here, we have examined whether ER binding sites that are located distal from the transcription start sites of estrogen target genes are functionally relevant. Similar to ER binding sites near the proximal promoter region, ER binding sites located at distal locations are occupied by ERs after estrogen stimulation. And, like proximal bound ERs, ERs occupied at distal sites can recruit coactivators and the RNA polymerase transcription machinery and mediate specific structural changes to chromatin. Furthermore, ERs occupied at the distal sites are capable of communicating with ERs bound at the promoter region, possibly via long range chromosome looping. In functional analysis, disruption of the response elements in the distal ER binding sites abrogated ER binding and significantly reduced transcriptional response. Finally, sequence comparison of the response elements at the distal sites suggests a high level of conservation across different species. Together, our data indicate that distal ER binding sites are bona fide transcriptional enhancers that are involved in long range chromosomal interaction, transcription complex formation, and distinct structural modifications of chromatin across large genomic spans.
|Oxysterol and diabetes activate STAT3 and control endothelial expression of profilin-1 via OSBP1.|
Romeo, GR; Kazlauskas, A
The Journal of biological chemistry 283 9595-605 2008
Endothelial dysfunction plays a central role in diabetic vascular disease, but its molecular bases are not completely defined. We showed previously that the actin-binding protein proflin-1 was increased in the diabetic endothelium and that attenuated expression of profilin-1 protected against atherosclerosis. Also 7-ketocholesterol up-regulated profilin-1 in endothelial cells via transcriptional mechanisms. The present study addressed the pathways responsible for profilin-1 gene expression in 7-ketocholesterol-stimulated endothelial cells and in the diabetic aorta. In luciferase reporter assays, the response to 7-ketocholesterol within the 5'-flanking region of profilin-1 was dependent on a single STAT response element. In aortic endothelial cells, 7-ketocholesterol enhanced STAT3 activation, which required JAK2 and tyrosine 394 phosphorylation of oxysterol-binding protein-1. These changes were recapitulated in the aorta of diabetic rats. Also 7-ketocholesterol in cultured endothelial cells and diabetes in the aorta elicited the recruitment of STAT3 and relevant coregulatory factors to the oxysterol-responsive region of the profilin-1 promoter. These events were required for profilin-1 up-regulation. These studies identify a previously unrecognized oxysterol-binding protein-mediated mode of activation of STAT3 that controls the expression of the proatherogenic protein profilin-1 in response to 7-ketocholesterol and the diabetic milieu.
|Estrogen receptor beta isoform-specific induction of transforming growth factor beta-inducible early gene-1 in human osteoblast cells: an essential role for the activation function 1 domain.|
Hawse, JR; Subramaniam, M; Monroe, DG; Hemmingsen, AH; Ingle, JN; Khosla, S; Oursler, MJ; Spelsberg, TC
Molecular endocrinology (Baltimore, Md.) 22 1579-95 2008
The estrogen receptors (ER) alpha and beta are important ligand-mediated transcription factors known to play significant biological roles in numerous tissues including bone. Despite the high homology shared by these receptors, recent studies have suggested that their function is largely unique. Although these receptors have been studied in detail for more than a decade, little data exist concerning the mechanisms by which these two proteins regulate distinct sets of genes. Using the TGFbeta-inducible early gene-1 (TIEG) as a model, we demonstrate that TIEG is rapidly induced in response to estrogen in osteoblasts by ERbeta, but not ERalpha. We have identified the regulatory elements utilized by ERbeta and have demonstrated that ERbeta recruits steroid receptor coactivator (SRC)1 and SRC2 to this regulatory region. Additionally, deletion of the ERbeta-activation function 1 (AF1) domain drastically decreases the estrogen induction of TIEG. Through the use of chimeric receptors, we have demonstrated that the AF1 domain of ERbeta is responsible for recruiting SRC1 and SRC2 and inducing the expression of TIEG in osteoblasts. Finally, SRC1, but not SRC2, is essential for TIEG induction by ERbeta. Overall, these data demonstrate that the estrogen induction of TIEG is ERbeta specific and that the AF1 domain of ERbeta confers this specificity. Finally, a novel and important role for ERbeta's AF1 is implicated in the recruitment of specific coactivators, suggesting that the AF1 may play a significant role in conferring the differences in regulation of gene expression by these two receptors.
|Progressive loss of estrogen receptor alpha cofactor recruitment in endocrine resistance.|
Naughton, C; MacLeod, K; Kuske, B; Clarke, R; Cameron, DA; Langdon, SP
Molecular endocrinology (Baltimore, Md.) 21 2615-26 2007
Differential expression of estrogen receptor-alpha (ERalpha) cofactors has been implicated in endocrine resistance in breast cancer. Using a three-stage MCF-7 cell-based model that emulates the clinical manifestation of acquired endocrine resistant breast cancer we now show, using a combination of chromatin immunoprecipitation and RNA interference, that there is a progressive loss of ERalpha cofactor recruitment to the estrogen-dependent pS2 gene and reduced requirement for cofactor expression. Maximal estrogen induced pS2 induction requires ERalpha and cofactor recruitment in MCF-7 cells, but in the progression to endocrine resistance these requirements are altered and expression has become less dependent on cofactors. Additionally, in estrogen-resistant MCF-7 cells there is a global loss of requirement of individual cofactors for proliferative cell growth indicating that other genes have lost the need for transcriptional cofactors. This loss of the requirement for cofactors may represent an important mechanism for gene misregulation in cancer.
|Estrogen receptor isoform-specific regulation of the retinoblastoma-binding protein 1 (RBBP1) gene: roles of AF1 and enhancer elements.|
Monroe, DG; Secreto, FJ; Hawse, JR; Subramaniam, M; Khosla, S; Spelsberg, TC
The Journal of biological chemistry 281 28596-604 2006
Estrogen (E2) is involved in mediating many important functions relevant to osteoblast biology through the actions of the estrogen receptors (ER) alpha and beta. To further understand the mechanisms of ER-specific regulation, we used microarray and reverse transcription-PCR analyses of E2-treated U2OS-ERalpha or -ERbeta cells and identified retinoblastoma-binding protein 1 (RBBP1) as a major E2-regulated gene. RBBP1 is a retinoblastoma cofactor involved in the control of osteoblastic proliferation. Although RBBP1 mRNA levels rapidly increased after 2 h of E2 treatment in both U2OS-ER-expressing lines, a sustained induction was only observed in U2OS-ERalpha cells. Examination of the RBBP1 genomic sequence revealed an ER response element and a Sp1 site located within the first intron. Chromatin immunoprecipitation analyses demonstrated that E2-dependent ERalpha binding to the intron 1 enhancer region was constitutive, whereas ERbeta binding was transient, consistent with the mRNA time course. Interestingly, transient transfection and receptor mutational studies revealed that RBBP1 induction by ERalpha only requires the Sp1 site, whereas ERbeta utilizes both the Sp1 and estrogen response elements binding sites for maximal E2-dependent activation. Stable U2OS transfectants containing a deletion of the ERalpha activation function 1 (AF1) resulted in a temporal mRNA induction profile similar to that of wild type ERbeta. Further, overexpression and chromatin immunoprecipitation analyses also demonstrated that E2-dependent RBBP1 induction is SRC2-dependent for both ER isoforms. These results describe an E2-dependent, ER isoform-specific transcriptional activation of the RBBP1 gene, which in part, is explained by the differential activity of ER AF1 and enhancer element binding.
|Steroid receptor coactivator-3 is required for progesterone receptor trans-activation of target genes in response to gonadotropin-releasing hormone treatment of pituitary cells.|
An, BS; Selva, DM; Hammond, GL; Rivero-Muller, A; Rahman, N; Leung, PC
The Journal of biological chemistry 281 20817-24 2006
Regulation of gonadotropin production involves interplay between steroids and neuropeptides, and we have examined the effects of gonadotropin-releasing hormones (GnRH I and GnRH II) on progesterone receptor (PR) activation in alphaT3-1 pituitary cells. Treatment with GnRHs activated a progester-one response element (PRE)-luciferase reporter gene, and this was blocked by protein kinase C and protein kinase A inhibitors but not by RU486. Treatment with GnRHs phosphorylated the PR at Ser(294) and increased PR translocation to the nucleus within 1 h. Interactions between the PR and several coactivators were examined, and treatment with GnRHs specifically induced PR-steroid receptor coactivator-3 (SRC-3) interactions within 8 h. In chromatin immunoprecipitation assays, recruitment of PR and SRC-3 by the PREs of the luciferase reporter gene or the gonadotopin alpha-subunit gene promoter was also increased by GnRHs within 8 h, while progesterone-induced recruitment of PR to the PREs occurred in association with much less SRC-3. A small interfering RNA knockdown of type I GnRH receptor levels reduced PR activation by GnRHs, while progesterone-dependent PR activation was unaffected. Moreover, small interfering RNA knockdown of SRC-3 abolished PRE-luciferase trans-activation by the PR in response to GnRHs. Collectively, these data indicate that PR activation by GnRHs in alphaT3-1 cells is type I GnRH receptor-mediated and that trans-activation of PR-responsive genes requires SRC-3 in this context.