Histone methyltransferase that specifically trimethylates 'Lys-9' of histone H3. H3 'Lys-9' trimethylation represents a specific tag for epigenetic transcriptional repression by recruiting HP1 (CBX1, CBX3 and/or CBX5) proteins to methylated histones. Mainly functions in euchromatin regions, thereby playing a central role in the silencing of euchromatic genes. H3 'Lys-9' trimethylation is coordinated with DNA methylation. Probably forms a complex with MBD1 and ATF7IP that represses transcription and couples DNA methylation and histone 'Lys-9' trimethylation. Its activity is dependent on MBD1 and is heritably maintained through DNA replication by being recruited by CAF-1. SETDB1 is targeted to histone H3 by TRIM28/TIF1B, a factor recruited by KRAB zinc-finger proteins.
Widely expressed. High expression in testis.
Belongs to the histone-lysine methyltransferase family. Suvar3-9 subfamily. Contains 1 MBD (methyl-CpG-binding) domain. Contains 1 post-SET domain. Contains 1 pre-SET domain. Contains 1 SET domain. Contains 2 Tudor domains.
The pre-SET, SET and post-SET domains are all required for methyltransferase activity. The 347-amino-acid insertion in the SET domain has no effect on the catalytic activity. Isoform 2 lacks all domains required for histone methyltransferase activity.
Nucleus. Chromosome. Associated with non-pericentromeric regions of chromatin. Excluded from nucleoli and islands of condensed chromatin.
Predicted band size : 155 kDa Samples: Whole cell lysate (10 mcg for 3 and 4; 50 mcg for 1 and 2) from 293T cells that were mock transfected (1 and 2) or transfected with a SETDB1 expression construct (3 and 4).
ab12317 (4ug/ml) staining KMT1E in human testis using an automated system (DAKO Autostainer Plus). Using this protocol there is strong staining of KMT1E in nuclear compartments of the germinal epithelium. Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer citrate pH6.1 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
Immunoprecipitation - SETDB1 antibody (ab12317)
Whole cell lysate (2 mg) from 293T cells that were mock transfected (1 and 2) or transfected with a SETDB1 expression construct (3 and 4). Rabbit anti-SETDB1 antibodies; ab12317 (2 and 4) and a competitor's antiserum (1 and 3) were used at 1 mcg/mg lysate for IP. ab12317 was used at 0.2 mcg/ml for Western Blot. Detection: Chemiluminescence with exposure times of 1 to 5 minutes.