An experimental assay was developed to search for proteins capable of antagonizing histone H1‐mediated general repression of transcription. T7 RNA polymerase templates containing an upstream scaffold‐associated region (SAR) were highly selectively repressed by H1 relative to non‐SAR control templates. This is due to the nucleation of H1 assembly into flanking DNA brought about by the numerous A‐tracts (AT‐rich sequences containing short homopolymeric runs of dA.dT base pairs) of the SAR. Partial, selective titration of these A‐tracts by the high mobility group (HMG) protein HMG‐I/Y led to the complete derepression of transcription from the SAR template by inducing the redistribution of H1 on to non‐SAR templates. SARs are associated with many highly transcribed regulated genes where they may serve to facilitate the HMG‐I/Y‐mediated displacement of histone H1 in chromatin. Indeed, HMG‐I/Y was found to be strongly enriched in the H1‐depleted subfraction which can be isolated from chromatin.