Functional Analysis of Ing1 and Ing4 in Cell Growth and Tumorigenesis: a Dissertation

Abstract

The five member Inhibitor of Growth (ING) gene family has been proposed to participate in the regulation of cell growth, DNA repair, inflammation, chromatin remodeling, and tumor suppression. All ING proteins contain a PHD motif implicated in binding to methylated histones and are components of large chromatin remodeling complexes containing histone acetyltransferase (HAT) and histone deacetylase (HDAC) enzymes, suggesting a role for ING proteins in regulating gene transcription. Additionally, forced overexpression studies performed in vitro have indicated that several ING proteins can interact with the p53 tumor suppressor protein and/or the NF-кB protein complex. Since these two proteins play well-established roles in numerous biological processes, several models have been proposed in the literature that ING proteins act as key regulators of cell growth and tumor suppression not only through their ability to modify gene transcription but also through their ability to alter p53 and NF-кB activity. However, these models have yet to be substantiated by in vivo experimentation. Research described in this dissertation utilizes a genetic approach to analyze the functional role of two ING proteins, Ing1b and Ing4, in regulating cell growth, inflammation, and tumorigenesis. Loss of p37Ing1b increased proliferation and DNA damage-induced apoptosis irrespective of p53 status in primary cells and mice. However, all other p53 responses were unperturbed. Additionally, p37Ing1b suppressed the formation of spontaneous follicular B-cell lymphomas in mice. Analysis of B-cells from these mice indicates that p37Ing1b inhibits the proliferation of B cells regardless of p53 status, and loss of p53 greatly accelerates the rate of B-cell lymphomagenesis in p37Ing1b-null mice, with double null mice presenting with aggressive diffuse large B-cell lymphomas (DLBL). Marker gene analysis in p37Ing1b/p53 null tumors indicates that these mice develop both non-germinal center and germinal center B cell-like DLBL, and also documents upregulation of NF-кB activity in both B-cells and tumors. Similarly, Ing4 -/- mice did not have altered p53 growth arrest or apoptosis, and did not develop spontaneous tumors. However, Ing4 -/- cells displayed reduced proliferation, and Ing4 -/- mice and macrophages were hypersensitive to treatment with LPS and exhibited decreased IкB gene expression and increased NF-кB activity. These studies demonstrate that Ing proteins can function to suppress spontaneous tumorigenesis and/or inflammatory responses without altering p53 activity, and identifies NF-кB as a biologically-relevant in vivo target of Ing1 and Ing4 signaling.