Figure 5. Effect of histone H1 subtypes on ATP-dependent chromatin remodeling.

MMTV minichromosomes were reconstituted in the presence of the histone H1 subtypes, as indicated, purified, resuspended in a 60 mM KCl and 5 mM MgCl₂ containing buffer, and incubated with Fok I that cleaves in nucleosome A and B of the MMTV promoter. Increasing nM concentrations of SWI/SNF (A) and NURF (B) complexes were added and incubated for 30 minutes at 30°C. The reaction was stopped and a linear extension PCR with a radiolabeled oligonucleotide was performed. The cleavage products were visualised in a 10% polyacrylamide denaturing gel and quantified using Image Quant software (Amersham). Minichromosomes of two independent purifications were tested and the results represented with the corresponding S.E.M in the graphs bellow. Abbreviations: Nuc A, nucleosome A; Nuc B, nucleosome B; U, uncut material; C, cut material.