The E2A locus is involved in chromosomal translocations associated with acute lymphoblastic leukemia. The most common of these involves a translocation between chromosomes 1 and 19 (t1;19), resulting in expression of the chimeric oncoprotein E2A-PBX1. A direct interaction between transcriptional activation domain 1 (AD1) of E2A and KIX domain of the histone acetyltransferase (HAT) /co-activator CBP is required for E2A-PBX1-mediated leukemia induction in mice. This thesis examines the functional consequences of the direct, physical interaction between E2A and CBP, for both proteins.
We demonstrate that the interaction between E2A and CBP/p300, as well as another HAT/co-activator, p/CAF, results in acetylation of E2A. Mutagenesis-based mapping studies identify several lysine residues as substrates for acetylation. Of particular interest, a conserved lysine (K34) located within AD1 is acetylated in vitro and in vivo. Substitution of this residue to arginine impairs transcriptional activation of a luciferase reporter while substitution to glutamine, mimicking the acetylation, restores E2A-mediated transcriptional activation.
Recent studies have shown that several transcription factors can modulate the intrinsic HAT activity of CBP/p300. We were surprised to find that E2A proteins enhance acetylation of histones by CBP, in vitro and in vivo, in a KIX domain-independent manner. Acetylation of E2A is also not required for stimulation of CBP/p300 histone acetylation. It appears that E2A interacts with the other CBP domains to mediate this effect, presumably through allosteric effects.
In summary, we demonstrate that acetylation of E2A plays a role in mediating the transcriptional activation activity of E2A. Furthermore, acetylation of E2A enhances its interaction with CBP/p300, at least in the presence of additional nuclear factors. We show evidence that p/CAF may mediate this effect. Enhancement of CBP/p300 HAT activity by oncogenic E2A-PBX1 proteins in vivo, suggests that some of its leukemia-promoting effects may be due to E2A-induced gain of function effects on CBP/p300. The enhanced interaction between acetylated E2A and CBP/p300, as well as the E2A-mediated stimulation of histone acetyltransferase activity might play a role in the DNA-binding-independent induction of proliferation. / Thesis (Ph.D, Pathology & Molecular Medicine) -- Queen's University, 2007-09-26 13:37:21.905
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:OKQ.1974/747 |
| Date | 01 October 2007 |
| Creators | Hyndman, Brandy Dawn |
| Contributors | Queen's University (Kingston, Ont.). Theses (Queen's University (Kingston, Ont.)) |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English, English |
| Detected Language | English |
| Type | Thesis |
| Format | 2459724 bytes, application/pdf |
| Rights | This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner. |
| Relation | Canadian theses |
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