Detection of Human SREBP2 by Western Blot.
Western blot shows lysates of HeLa human cervical epithelial carcinoma cell line and Raji human Burkitt's lymphoma cell line. Gels were loaded with 15 µg of nuclear extracts (Nuc). PVDF membrane was probed with 2 µg/mL of Goat Anti-Human SREBP2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7119) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). Specific bands were detected for SREBP2 at approximately 125 kDa and 60-70 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
Detection of SREBP2-regulated Genes by Chromatin Immunoprecipitation.
HUVEC human umbilical vein endothelial cells were serum-starved for 5 hours, fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. SREBP2/DNA complexes were immunoprecipitated using 5 μg Goat Anti-Human SREBP2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7119) or control antibody (Catalog # AB-108-C) for 15 minutes in an ultrasonic bath, followed by Biotinylated Anti-Goat IgG Secondary Antibody (Catalog # BAF109). Immunocomplexes were captured using 50 μL of MagCellect Streptavidin Ferrofluid (Catalog # MAG999) and DNA was purified using chelating resin solution. The ABCA1 promoter was detected by standard PCR. In the figure, "Input" represents total cell lysate DNA.
SREBP2 in HeLa Human Cell Line.
SREBP2 was detected in immersion fixed HeLa human cervical epithelial carcinoma cell line using Goat Anti-Human SREBP2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7119) at 15 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # NL001) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
Preparation and Storage
Sterile PBS to a final concentration of 0.2 mg/mL.
Reconstitution Buffer Available
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.
SREBP2 (Sterol Regulatory Element-Binding Protein 2; also bHLHD2 and SREBF2) is a 120-125 kDa member of the SREBP family of proteins. It is ubiquitously expressed and found in the intracellular membrane fraction of cells. SREBP2 is a transcriptional factor initially embedded in the ER as an inactive precursor associated with SCAP. When necessary, SCAP mediates SREBP2 transfer to the Golgi, where two resident proteases remove the N-terminus from SREBP2, and the N-terminus is transported into the nucleus. Here, SREBP2 acts as a transcription factor, activating the LDLR and cholesterol synthesis genes. The human SREBP2 precursor is an 1141 amino acid (aa) two transmembrane protein whose N- and C-termini are cytoplasmic. The two cytoplasmic domains span aa 1-479 and 555-1141, respectively. Proteolytic cleavage between Leu484-Cys485 generates the 64-66 kDa SREBP2 transcription factor. This fragment contains a bHLH DNA binding domain (aa 330-380) and one Leu zipper region (aa 381-401). Homodimerization of SREBP2 is necessary for nuclear translocation. There is one potential isoform that shows a deletion of aa 274-276 coupled to a 96 aa substitution for aa 580-1141. A second isoform (known in rodent) shows a premature truncation after Val463 and runs at 55 kDa in SDS-PAGE. Over aa 242-450, human SREBP2 shares 97% aa identity with mouse SREBP2.
Sterol Regulatory Element Binding Transcription Factor 2
Entrez Gene IDs:
6721 (Human); 20788 (Mouse); 300095 (Rat)
bHLhd2; bHLHd2sterol regulatory element-binding protein 2; Class D basic helix-loop-helix protein 2; SREBF2; SREBP2; SREBP2SREBP-2; sterol regulatory element binding transcription factor 2; Sterol regulatory element-binding transcription factor 2
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