Mechanisms which control the stable silencing of viral and cellular genes: Coordination of histone de-acetylation, methylation and heterochromatin formation by KAP-1

Frank J. Rauscher, III, PhD

The Wistar Institute, Philadelphia, PA, USA

Gene regulation at the level of transcriptional initiation is critical for development and homeostasis in higher vertebrates. Deregulation of coordinated gene expression underlies many viral and cellular pathogenic states. A great emphasis has been placed on defining the exact molecular mechanisms whereby sets of genes are selected for coordinated activation or repression and how these, once initiated, are maintained. The past few years has seen an explosion of research on the regulation of chromatin structure as a primary mechanism of transcriptional control. In particular, the post-translational modification of core histone tails via phosphorylation, acetylation/de-acetylation, methylation and ubiquitination has emerged as a key event. Multi-subunit macromolecular complexes have been isolated and shown to contain these activities while genetic studies have proven their requirement for promotor-specific gene regulation. However, much less clear is how these complexes are recruited to promotors in vivo, and even more enigmatic is how their enzymatic activities are spatially and temporarily coordinated. We have been studying a transcriptional co-repressor protein, KAP-1, which has the capability to both bind to sequence-specific DNA binding subunits (KRAB domain-containing Zinc Finger proteins) and recruit complexes containing histone de-acetylase (NuRD), histone methylases (KIS-1) and heterochromatin proteins (HP1s). The KAP1 co-repressor contains at least four distinct surfaces for associating with these activities and may function to sequentially recruit them to target promoters in vivo. A combination of CHIP, enzyme accessibility and Immuno-FISH assays have confirmed that endogenous genes silenced by the KAP-1 system display: 1) a closed chromatin confirmation 2) a compliment of core histones containing the predicted modifications, and 3) a direct association with the heterochromatin protein HP1 and condensed chromatin. These results form a paradigm for understanding the coordination of chromatin modifications that lead to stable silencing of gene expression. Our current studies on the regulation of this system will be presented.

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Molecular Basis of Oncogenesis.