Mammalian GCN5 and P/CAF Acetyltransferases Have Homologous Amino-Terminal Domains Important for Recognition of Nucleosomal Substrates

ABSTRACT

The yeast transcriptional adapter Gcn5p serves as a histone acetyltransferase, directly linking chromatin modification to transcriptional regulation. Two human homologs of Gcn5p have been reported previously, hsGCN5 and hsP/CAF (p300/CREB binding protein [CBP]-associated factor). While hsGCN5 was predicted to be close to the size of the yeast acetyltransferase, hsP/CAF contained an additional 356 amino-terminal residues of unknown function. Surprisingly, we have found that in mouse, both the GCN5and the P/CAF genes encode proteins containing this extended amino-terminal domain. Moreover, while a shorter version of GCN5 might be generated upon alternative or incomplete splicing of a longer transcript, mRNAs encoding the longer protein are much more prevalent in both mouse and human cells, and larger proteins are detected by GCN5-specific antisera in both mouse and human cell extracts. Mouse GCN5 (mmGCN5) and mmP/CAF genes are ubiquitously expressed, but maximum expression levels are found in different, complementary sets of tissues. Both mmP/CAF and mmGCN5 interact with CBP/p300. Interestingly,mmGCN5 maps to chromosome 11 and cosegregates withBRCA1, and mmP/CAF maps to a central region of chromosome 17. As expected, recombinant mmGCN5 and mmP/CAF both exhibit histone acetyltransferase activity in vitro with similar substrate specificities. However, in contrast to yeast Gcn5p and the previously reported shorter form of hsGCN5, mmGCN5 readily acetylates nucleosomal substrates as well as free core histones. Thus, the unique amino-terminal domains of mammalian P/CAF and GCN5 may provide additional functions important to recognition of chromatin substrates and the regulation of gene expression.

ACKNOWLEDGMENTS

We thank Jerry Workman and Patrick Grant for the kind gift of HeLa cell mononucleosomes and histones. We thank David Allis for the gift of H3 amino-terminal peptides, and we thank E. Smith and D. Allis for the gift of oligomers for PCR and for sharing results prior to publication. We also thank Shelley Berger for antiserum specific for hsGCN5, Yongshen Ren for the gift of HeLa cell nuclear extracts, and Yoshihiro Nakatani for the GST-CBP and GST-p300 fusion constructs and hsP/CAF antibodies. Some DNA sequencing was performed by the UTMDACC Sequencing Core Facility. We are grateful to Lucy Rowe and Mary Barter at the Jackson Laboratory for their assistance in the mouse chromosome mapping analysis. We thank Karen Hensley for help in preparation of some graphics and Aurora Diaz for help in preparing the manuscript.

W.X. is supported by a Rosalie B. Hite Fellowship, and D.G.E. is supported by a Theodore Law UCF Scientific Fund Fellowship. This work was supported by grants to S.Y.R. from the Robert A. Welch Foundation, the USARMC, and the Breast Cancer Research Center at UTMDACC.

The first two authors contributed equally to this work.