Chromatin antibody

Kretz-Rommel A Rubin RL (1999) Persistence of autoreactive T cell drive is required to elicit anti-chromatin antibodies in a murine model of drug-induced lupus. J Immunol, 162(2) 813-820. 

Binding of nitroso-procainamide to histone proteins may perturb chromatin structure or catabolism, resulting in immunogenic forms of DNA-free histones. In fact, all sera of patients (n = 24) with procainamide-induced Lupus showed IgG and IgM antibody activity against various histone components of chromatin (chromosome subunits)122. The nature of the procainamide adduct to histone proteins still awaits elucidation. 

The first indication that modification of specific tail residues were linked to chromatin functional states, came from immunostaining of Drosophila polytene chromosomes with antibodies specific for H4 acetylated at defined lysines [13]. As shown in Fig. 2A, H4 acetylated at lysine 16 (H4acK16) was found almost exclusively on the transcriptional hyperactive male X chromosome (Fig. 2). (Genes on the Drosophila male X are transcribed twice as fast as their female counterparts so as to equalize levels of X-linked gene products between XY males and XX females.) In addition, H4 lysine 12 was found to remain acetylated in centric heterochromatin, while lysines 5, 8, and 16 were all under-acetylated [13]. These observations led to the suggestion that the histone N-terminal tails constitute nucleosome surface markers that can be recognized by non-histone proteins in a modification-dependent manner to alter the functional state of chromatin [13]. 

Jeppesen, P., Mitchell, A., Turner, B.M., and Perry, P. (1991) Antibodies to defined histone epitopes reveal variations in chromatin conformation and underacetylation of centric heterochromatin in human metaphase chromosomes. Chromosoma 100, 322-332. 

The basic principle behind the ChIP method is relatively simple It is based on the selective enrichment of a chromatin fraction containing a specific antigen (e.g. transcription factors, DNA binding proteins, modified histones, etc.) by an immunoprecipitation step. Specific (important, see below ) antibodies that recognize a protein of interest or the modified form of a protein can be used to determine the relative abundance of it within DNA regions. 

For re-ChIP (or double ChIP), two sequential rounds of immunoprecipitations are performed using two different antibodies. Re-ChIP allows one to determine whether two antigens occur together on the same chromatin fragment or on different fragments and can be used to assemble proteins in complexes [23]. 

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