Histone Acetyltransferase HBO1 Interacts with the ORC1 Subunit of the Human Initiator Protein

Masayoshi Iizuka,Bruce Stillman

doi:10.1074/jbc.274.33.23027

Masayoshi Iizuka, Bruce Stillman

Open Access

https://doi.org/10.1074/jbc.274.33.23027

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Abstract

The origin recognition complex (ORC) is an initiator protein for DNA replication, but also effects transcriptional silencing in Saccharomyces cerevisiae and heterochromatin function in Drosophila. It is not known, however, whether any of these functions of ORC is conserved in mammals. We report the identification of a novel protein, HBO1 (histone acetyltransferase binding to ORC), that interacts with human ORC1 protein, the largest subunit of ORC. HBO1 exists as part of a multisubunit complex that possesses histone H3 and H4 acetyltransferase activities. A fraction of the relatively abundant HBO1 protein associates with ORC1 in human cell extracts. HBO1 is a member of the MYST domain family that includes S. cerevisiae Sas2p, a protein involved in control of transcriptional silencing that also has been genetically linked to ORC function. Thus the interaction between ORC and a MYST domain acetyltransferase is widely conserved. We suggest roles for ORC-mediated acetylation of chromatin in control of both DNA replication and gene expression.

Highlights

The origin recognition complex (ORC)1 is a key protein for the initiation of DNA replication in eukaryotes
No in-frame stop codon was found upstream of the first methionine, this methionine is likely to be the initiation codon, because the size of the protein translated from this cDNA is the same as that of the protein present in human cells
Based upon our finding that the L74 cDNA encodes a protein that binds to human ORC1 protein (hORC1) and is a subunit of a histone acetyltransferase, we termed the protein Histone acetyltransferase Bound to ORC 1 (HBO1), an acronym for Histone acetyltransferase Binding to hORC1

Summary

EXPERIMENTAL PROCEDURES

Identification of a cDNA Encoding a Human ORC1 Protein-binding Protein—Human full-length ORC1 cDNA from pKG28 (11) was cloned into the SmaI and BamHI sites of the pBTM116 vector (a gift from Linda van Aelst, Cold Spring Harbor Laboratory), carrying the TRP1 gene and a lexA DNA-binding domain, to generate plexAhORC1. Yeast strain L40 (36, 37) (MAT a his3⌬200 trp901 leu 112 ade lys2– 801am URA3::(lexAop)8-lacZ LYS2::(lexAop)8-HIS3) harboring the plexAhORC1 plasmid was transformed with a cDNA library cloned in the pGADGH vector (a gift from Linda van Aelst), carrying the LEU2 gene and the GAL4 activation domain. Restoration of ␤-galactosidase activity was assayed to test the specificity of interaction after adding back the plasmid plexAhORC1 or plexAHRAS by mating with AMR70

ORC Interacting Protein

RESULTS AND DISCUSSION

Masayoshi Iizuka and Bruce Stillman