Product Pathways - Nuclear Receptor Signaling

• pathway
• application references
• companion products
• datasheet PDF
• MSDS PDF
• protocols

PPARγ Regulated Fatty Acid Metabolism Antibody Sampler Kit #8660

Kit Includes	Quantity	Applications	Reactivity	MW (kDa)	Isotype
Phospho-AMPKα (Thr172) (40H9) Rabbit mAb #2535	40 µl	W IP IHC-P	H M R Hm Mk Dm Sc (C) (Z) (B) (Pg)	62	Rabbit IgG
AMPKα (23A3) Rabbit mAb #2603	40 µl	W	H M R Mk	62	Rabbit IgG
CBP (D6C5) Rabbit mAb #7389	40 µl	W IP IF-IC ChIP	H M R Mk	300	Rabbit IgG
GCN5L2 (C26A10) Rabbit mAb #3305	40 µl	W IP IF-IC	H M R Mk (B) (Hr)	94	Rabbit IgG
PPARγ (C26H12) Rabbit mAb #2435	40 µl	W IHC-P IF-IC	H M (R)	53, 57	Rabbit IgG
SirT1 (C14H4) Rabbit mAb #2496	40 µl	W IP	H	120	Rabbit
RXRα Antibody #5388	40 µl	W IP	H R Mk	53	Rabbit
Anti-rabbit IgG, HRP-linked Antibody #7074	100 µl				Goat

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IF-IC=Immunofluorescence (Immunocytochemistry)  ChIP=Chromatin IP
Reactivity Key:  H=Human  M=Mouse  R=Rat  Hm=Hamster  Mk=Monkey  C=Chicken  Dm=D. melanogaster  Z=Zebrafish  B=Bovine  Pg=Pig  Sc=S. cerevisiae  Hr=Horse
Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Specificity / Sensitivity

Phospho-AMPKα (Thr172) (40H9) Rabbit mAb detects endogenous AMPKα only when phosphorylated at Thr172. Phospho-AMPKα (Thr172) (40H9) Rabbit mAb detects both α1 and α2 isoforms of the catalytic subunit, but does not detect the regulatory β or γ subunits. AMPKα (23A3) Rabbit mAb, CBP (D6C5) Rabbit mAb, GCN5L2 (C26A10) Rabbit mAb, PPARγ (C26H12) Rabbit mAb, SirT1 (C14H4) Rabbit mAb, and RXRα Antibody all detect endogenous levels of their respective total proteins.

Description

PPARγ Regulated Fatty Acid Metabolism Antibody Sampler Kit provides an economical means to evaluate PPARγ and related proteins involved in lipid metabolism. This kit contains enough primary antibody to perform four western blots per primary.

Source / Purification

Monoclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Thr172 of human AMPKα protein or with a synthetic peptide corresponding to the respective sequences of human AMPKα, CBP, GCN5L2, PPARγ, and SirT1. Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human RXRα protein. Antibodies are purified by protein A and peptide affinity chromatography.

Background

AMPK is a heterotrimeric complex composed of a catalytic α subunit and regulatory β and γ subunits, each of which is encoded by two or three distinct genes (α1, 2; β1, 2; γ1, 2, 3) (1). The kinase is activated by an elevated AMP/ATP ratio due to cellular and environmental stress, such as heat shock, hypoxia, and ischemia (1). The tumor suppressor LKB1 phosphorylates AMPKα at Thr172 in the activation loop, and this phosphorylation is required for AMPK activation (2-4). Accumulating evidence indicates that AMPK not only regulates the metabolism of fatty acids and glycogen, but also modulates protein synthesis and cell growth through EF2 and TSC2/mTOR pathways, as well as blood flow via eNOS/nNOS (5).CBP (CREB-binding protein) is a transcriptional co-activator that associates with PPARγ (6,7). CBP also contains histone acetyltransferase (HAT) activity, allowing it to acetylate histones and other proteins (7).General Control of Amino Acid Synthesis Yeast Homolog Like 2 (GCN5L2) is a transcription adaptor protein and a histone acetyltransferase (HAT) that functions as the catalytic subunit of the STAGA and TFTC transcription coactivator complexes (8). GCN5L2 is 73% homologous to the p300/CBP-associated factor PCAF, another HAT protein found in similar complexes (9). GCN5L2 acetylates non-histone proteins such as the transcription co-activator PGC1-α (10).Peroxisome proliferator-activated receptor γ (PPARγ) is a member of the ligand-activated nuclear receptor superfamily and functions as a transcriptional activator (11). PPARγ is preferentially expressed in adipocytes as well as in vascular smooth muscle cells and macrophage (12).The Silent Information Regulator (SIR2) family of genes is a highly conserved group of genes that encode nicotinamide adenine dinucleotide (NAD)-dependent protein deacetylases, also known as class III histone deacetylases (13). SirT1, the mammalian ortholog of Sir2, is a nuclear protein implicated in the regulation of many cellular processes, including apoptosis, cellular senescence, endocrine signaling, glucose homeostasis, aging, and longevity. Targets of SirT1 include PPARγ (14), and the PPARγ coactivator-1α (PGC-1α) protein (15). Deacetylation of PPARγ and PGC-1α regulates the gluconeogenic/glycolytic pathways in the liver and fat mobilization in white adipocytes in response to fasting (14,15).The human retinoid X receptors (RXRs) are type-II nuclear hormone receptors encoded by three distinct genes (RXRα, RXRβ, and RXRγ) and bind selectively and with high affinity to the vitamin A derivative, 9-cis-retinoic acid. Nuclear RXRs form heterodimers with PPAR to help regulate transcription during lipid metabolism (16).

1. Carling, D. (2004) Trends Biochem Sci 29, 18-24.
2. Hawley, S.A. et al. (1996) J Biol Chem 271, 27879-87.
3. Lizcano, J.M. et al. (2004) EMBO J 23, 833-43.
4. Shaw, R.J. et al. (2004) Proc Natl Acad Sci U S A 101, 3329-35.
5. Hardie, D.G. (2004) J Cell Sci 117, 5479-87.
6. Goodman, R.H. and Smolik, S. (2000) Genes Dev 14, 1553-77.
7. Chan, H.M. and La Thangue, N.B. (2001) J Cell Sci 114, 2363-73.
8. Candau, R. et al. (1996) Mol Cell Biol 16, 593-602.
9. Yang, X.J. et al. (1996) Nature 382, 319-24.
10. Lerin, C. et al. (2006) Cell Metab 3, 429-38.
11. Tontonoz, P. et al. (1995) Curr Opin Genet Dev 5, 571-6.
12. Rosen, E.D. et al. (1999) Mol Cell 4, 611-7.
13. Guarente, L. (1999) Nat Genet 23, 281-5.
14. Picard, F. et al. (2004) Nature 429, 771-6.
15. Rodgers, J.T. et al. (2005) Nature 434, 113-8.
16. Gronemeyer, H. et al. (2004) Nat Rev Drug Discov 3, 950-64.

Application References

Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!

Protocols

2435:

IHC / Paraffin, Immunofluorescence, Western Blotting

2496:

Immunoprecipitation, Western Blotting

2535:

IHC / Paraffin, Immunoprecipitation, Western Blotting

2603:

Western Blotting

3305:

Immunofluorescence, Immunoprecipitation, Western Blotting

5388:

Immunoprecipitation, Western Blotting

7389:

Immunofluorescence, Immunoprecipitation, Western Blotting

Companion Products

• 7071 Phototope^®-HRP Western Blot Detection System, Anti-rabbit IgG, HRP-linked Antibody
• 7003 20X LumiGLO^® Reagent and 20X Peroxide
• 9998 BSA
• 9999 Nonfat Dry Milk
• 7720 Prestained Protein Marker, Broad Range (Premixed Format)
• 7727 Biotinylated Protein Ladder Detection Pack
• 9997 Tris Buffered Saline with Tween 20 (TBST-10X)
• 7722 Blue Loading Buffer Pack
• 7723 Red Loading Buffer Pack
• 7074 Anti-rabbit IgG, HRP-linked Antibody
• 7075 Anti-biotin, HRP-linked Antibody

Selected rabbit monocloal antibodies are produced under license (granting certain rights including those under U. S. Patents No. 5,675,063 and in some instances 7,429,487) from Epitomics, Inc.

---

For Research Use Only. Not For Use In Diagnostic Procedures.