Off-campus UMass Amherst users: To download campus access dissertations, please use the following link to log into our proxy server with your UMass Amherst user name and password.

Non-UMass Amherst users: Please talk to your librarian about requesting this dissertation through interlibrary loan.

Dissertations that have an embargo placed on them will not be available to anyone until the embargo expires.

Author ORCID Identifier

N/A

AccessType

Open Access Dissertation

Document Type

dissertation

Degree Name

Doctor of Philosophy (PhD)

Degree Program

Animal Biotechnology & Biomedical Sciences

Year Degree Awarded

2014

Month Degree Awarded

February

First Advisor

Janice C. Telfer

Second Advisor

Cynthia Baldwin

Third Advisor

Wilmore Webley

Subject Categories

Animal Sciences | Biotechnology | Veterinary Medicine

Abstract

The transcription factors Runx1 and Runx3 are required for permanent silencing of CD4 in maturing CD8+ thymocytes. Runx binding to consensus sites within the CD4 silencer region is required for CD4 silencing post-positive selection. The Runx nuclear matrix targeting sequence (NMTS) is required for CD4 silencing and is implicated in binding to the histone acetyltransferase (HAT) p300, histone deacetylases (HDAC), and histone methyltransferase (HMT) SUV39H1 proteins. Epigenetic modifications of chromatin or post-translational modifications of Runx itself as a result of Runx association with these enzymes may be important for establishment of long-term CD4 silencing. In this study, we show that treatment of thymocytes with the HDAC inhibitor trichostatin A (TSA) and a resulting increase in histone acetylation in the CD4 silencer region results in an increase in CD4 silencing in a Runx-independent manner. We evaluated the role played by Runx lysines and their potential post-translational modification in CD4 silencing by mutational analysis of nine Runx lysine residues. Disruption of lysines within the Runt DNA binding domain known to reduce Runx DNA-binding activity resulted in CD4 derepression, indicating that Runx DNA-binding is required for CD4 silencing. Mutation of other lysines not involved with DNA-binding and reported to be acetylated or methylated did not affect CD4 silencing by Runx. The transduction of thymocytes with the C-terminally truncated Runx1 or Runx1 lacking the NMTS fused with the p300 HAT domain, HDAC1, or SUV39H1 resulted in CD4 derepression, indicating that enforced association of these individual enzymes with the Runx DNA-binding domain promotes CD4 transactivation rather than CD4 silencing. Profiling chromatin marks present under c-terminally truncated Runx1d.190 treatment conditions revealed H3K9me3/H3K4me3 coenrichment in CD4 promoter and silencer regions suggesting the involvement of a dynamic instruction profile in the establishment of CD4 silencing.

DOI

https://doi.org/10.7275/wjdk-3t13

Download

Share

COinS