#2616 HP1α Antibody

Western Blotting

Western blot analysis of lysates from HeLa, NIH/3T3, C6 and COS cells, using HP1alpha antibody.

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma, using HP1alpha Antibody in the presence of control peptide (left) or HP1 alpha blocking peptide #1004 (right).

IF-IC

Confocal immunofluorescent image of HeLa cells labeled with HP1α antibody (green). Actin filaments have been labeled with Alexa Fluor^® 555 phalloidin (red).

Flow Cytometry

Flow cytometric analysis of untreated HeLa cells, using HP1alpha antibody (blue) compared to a nonspecific negative control antibody (red).

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from mES cells and either HP1α Antibody #2616 or Normal Rabbit IgG #2729, using SimpleChIP^® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP^® Mouse MEST Intron 1 Primers #12870, SimpleChIP^® Mouse MEST Promoter Primers #12928, mouse Intracisternal A-Particle (IAP) LTR promoter primers, and mouse primers specific to a non-genic, non-repetitive region. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.

Heterochromatin protein 1 (HP1) is a family of heterochromatic adaptor molecules involved in both gene silencing and higher order chromatin structure (1). All three HP1 family members (α, β, and γ) are primarily associated with centromeric heterochromatin; however, HP1β and γ also localize to euchromatic sites in the genome (2,3). HP1 proteins are approximately 25 kDa in size and contain a conserved amino-terminal chromodomain, followed by a variable hinge region and a conserved carboxy-terminal chromoshadow domain. The chromodomain facilitates binding to histone H3 tri-methylated at Lys9, a histone "mark" closely associated with centromeric heterochromatin (4,5). The variable hinge region binds both RNA and DNA in a sequence-independent manner (6). The chromoshadow domain mediates the dimerization of HP1 proteins, in addition to binding multiple proteins implicated in gene silencing and heterochromatin formation, including the SUV39H histone methyltransferase, the DNMT1 and DNMT3a DNA methyltransferases, and the p150 subunit of chromatin-assembly factor-1 (CAF1) (7-9). In addition to contributing to heterochromatin formation and propagation, HP1 and SUV39H are also found complexed with retinoblastoma (Rb) and E2F6 proteins, both of which function to repress euchromatic gene transcription in quiescent cells (10,11). HP1 proteins are subject to multiple types of post-translational modifications, including phosphorylation, acetylation, methylation, ubiquitination, and sumoylation, suggesting multiple means of regulation (12-14).

1. Maison, C. and Almouzni, G. (2004) Nat. Rev. Mol. Cell Biol. 5, 296-304.
2. Minc, E. et al. (2000) Cytogenet. Cell Genet. 90, 279-284.
3. Nielsen, A.L. et al. (2001) Mol. Cell 7, 729-739.
4. Lachner, M. et al. (2001) Nature 410, 116-120.
5. Bannister, A.J. et al. (2001) Nature 410, 120-124.
6. Muchardt, C. et al. (2002) EMBO Rep. 3, 975-981.
7. Yamamoto, K. and Sonoda, M. (2003) Biochem. Biophys. Res. Commun. 301, 287-292.
8. Fuks, F. et al. (2003) Nucleic Acids Res. 31, 2305-2312.
9. Murzina, N. et al. (1999) Mol. Cell 4, 529-540.
10. Nielsen, S.J. et al. (2001) Nature 412, 561-565.
11. Ogawa, H. et al. (2002) Science 296, 1132-1136.
12. Minc, E. et al. (1999) Chromosoma 108, 220-234.
13. Zhao, T. et al. (2001) J. Biol. Chem. 276, 9512-9518.
14. Lomberk, G. et al. (2006) Nat. Cell Biol. 8, 407-415.