The initiation and pattern of spread of histone H4 acetylation parallel the order of transcriptional activation of genes in the aflatoxin cluster

The 27 genes involved in aflatoxin biosynthesis are clustered within a 70 kb region in the Aspergillus parasiticus genome. Using chromatin immunoprecipitation, we demonstrated a positive correlation between the initiation and spread of histone H4 acetylation in aflatoxin promoters and the onset of accumulation of aflatoxin proteins and aflatoxin. Histone H4 acetylation in the pksA (encodes an 'early' biosynthetic pathway enzyme) promoter peaked at 30 h, prior to the increased acetylation in the omtA and ordA (encode 'late' enzymes) promoters detected at 40 h. The specific order in which pksA, ver-1 (encodes a'middle'enzyme) and omtA transcripts accumulated in cells paralleled the pattern of spread of histone H4 acetylation. Binding of AflR, a positive regulator of aflatoxin bilosynthesis, to the ordA promoter showed a positive correlation with the spread of histone H4 acetylation. The data suggest that the order of genes within the aflatoxin cluster determines the timing and order of transcriptional activation, and that the site of initiation and spread of histone H4 acetylation mediate this process. Our data indicate that the aflatoxin and adjacent sugar utilization clusters are part of a larger 'regulatory unit'.