Characterization
Description(s):Transcriptional co-regulators interact specifically and non-covalently with one or multiple DNA-binding transcription factors (TFs) to either activate or repress the transcription of specific genes. Dysregulated expression of coregulator, such as SRC-1/NcoA-1, p300, CBP and HADC, has a significant effect on the regulation of a TF in the recruitment of other transcription factors and chromatin remodeling. For example, the upregulation of SRC-1 has known roles in mediating steroid receptor transcription in androgen receptor activity as well as prostate cancer progression.
Signosis has developed TF-Coregulator Interaction Plate Array Assay II allowing for high throughput studying of co-regulator interaction networks with 48 different TFs.
Applicable Grid:
| 1 | 2 | 3 |
4 |
5 |
6 |
7 | 8 | 9 | 10 |
11 |
12 | |
| A | AP1 | CDP | GATA |
NF-1 |
Pit |
Stat3 |
AP1 | CDP | GATA |
NF-1 |
Pit |
Stat3 |
| B | AP2 | CREB | GR/PR | NFAT | PPAR | Stat4 | AP2 | CREB | GR/PR | NFAT | PPAR | Stat4 |
| C | AR | E2F-1 |
HIF |
NF-E2 | PXR | Stat5 | AR | E2F-1 |
HIF |
NF-E2 | PXR | Stat5 |
| D | ATF2 | EGR | HNF4 | NFkB | SMAD | Stat6 | ATF2 | EGR | HNF4 | NFkB | SMAD | Stat6 |
| E | Brn-3 | ER | IRF | OCT4 | Sp1 | TCF/LEF | Brn-3 | ER | IRF | OCT4 | Sp1 | TCF/LEF |
| F | C\EBP | Ets | MEF2 | p53 | SRF | TR | C\EBP | Ets | MEF2 | p53 | SRF | TR |
| G | CAR | FAST-1 | Myb | Pax-5 | SATB1 | YY1 | CAR | FAST-1 | Myb | Pax-5 |
SATB1 |
YY1 |
| H | CBF | GAS/ISRE | Myc-Max | Pbx1 | Stat1 | TFIID | CBF | GAS/ISRE | Myc-Max | Pbx1 | Stat1 | TFIID |
Principle
Signosis’ TF-Coregulator Interaction Plate Array can simultaneously profile the transcriptional interaction of multiple TFs with a co-regulator of interest. In this assay, a series of unique biotin-labeled probes are provided that correspond with the consensus sequences of individual TF DNA-binding sites. Therefore, each probe represents an individual TF. When the probe mix is incubated with nuclear extract, individual probes bind to their corresponding TF. The co-regulator of interest is then immunoprecipitated, along with transcriptionally interacting TFs, using a corresponding antibody and protein G or A agarose beads in a tube. Unbound probes and proteins are washed away. The bound probes are then detached from the complex and are subsequently denatured. The biotin-labeled DNA strands are hybridized on a pre-coat plate and detected with streptavidin-HRP and substrate. The detected signals reflect the interacting TFs with the particular co-regulator of interest. Luminescence is reported as relative light units (RLUs) on a microplate luminometer.
