Leucine zipper motif of chicken histone acetyltransferase-1 is essential for in vivo and in vitro interactions with the p48 subunit of chicken chromatin assembly factor-1

We cloned cDNA encoding chicken cytoplasmic histone acetyltransferase-1, ch HAT-1, comprising 408 amino acids including a putative initiation Met. It e xhibits 80.4% identity to the human homolog and possesses a typical leucine zipper motif, The glutathione S-transferase (GST) pull-down assay, involvi ng truncated and missense mutants of the chicken chromatin assembly factor- 1 (chCAF-1)p48, revealed not only that a region (comprising amino acids 376 -405 of chCAF-1 p48 and containing the seventh WD dipeptide motif) binds to chHAT-1 in vitro, but also that mutation of the motif has no influence on the in vitro interaction. The GST pull-down assay, involving truncated and missense chHAT-1 mutants, established that a region, comprising amino acids 380-408 of chHAT-1 and containing the leucine zipper motif, is required fo r its in vitro interaction with chCAF-1p48. In addition, mutation of each o f four Leu residues in the leucine zipper motif prevents the in vitro inter action. The yeast two-hybrid assay revealed that all four Leu residues with in the leucine zipper motif of chHAT-1 are necessary for its in vivo intera ction with chCAF-1p48, These results indicate not only that the proper leuc ine zipper motif of chHAT-1 is essential for its interaction with chCAF-1p4 8, but also that the propeller structure of chCAF-1p48 expected to act as a platform for protein-protein interactions may not be necessary for this in teraction of chHAT-1.