A significant gap in the field of cancer research is the lack of unbiased, genome-wide studies describing how activated Notch regulates the genomes of cancer cells. We propose to begin to fill this gap by elucidating how activated Notchl (ICNI) interacts with the genomes of T-ALL cells to support leukemia cell growth. We will address this central issue in T-ALL pathogenesis by pursuing the following two specific aims:
Aim 1. To determine how Notchl transactivates target genes Current models for Notch regulation of target genes have been based on analyses of only a few genes (e.g., i-ies1) in diverse cellular contexts. Our preliminary data suggest that individual Notchl binding sites show differential sensitivity to y-secretase inhibitors (GSls), a characteristic that may define dynamic NREs and which points to an unexpected pool of GSI-refractory chromatin-associated Notchl. In this aim, we will first use ChlP-Seq to determine the GSI-sensitivity of chromatin-associated Notchl on a genome-wide basis. Then, with a full set of robust target genes and GSI-sensitive Notchl binding sites in hand, we will test and refine models of Notchl regulation of gene expression in T-ALL cells. We anticipate that these studies will illuminate fundamental mechanisms of target gene regulation by Notch and reveal new opportunities for therapeutic targeting ofthe Notch pathway.
Aim 2. To determine the contributions of cis-regulatory co-factors to Notchl target gene expression Motif analysis carried out in human and murine T-ALL cells showed that Notchl genomic binding sites are often flanked by motifs for Runx factors or Ets factors, or overlap precisely with binding sites for Znf143. In preliminary data, we have observed that Runx factors and Ets factors appear to co-regulate important Notchl target genes such as iL7R that contribute to the growth and survival of T-ALL cells. We will expand upon these data to study how Ets and Runx factors co-regulate key target genes, and will also elucidate the pervasive, but complex relationship of Notchl and Znf143. Together, the work described in these two aims will substantially advance our understanding of the molecular pathogenesis of T-ALL, and in doing so create new opportunities for targeting of the Notch pathway in cancer and in other forms of disease related to dysregulated Notch signaling.

Public Health Relevance

; Notch transforms cells by activating gene expression, yet little is know about details of how this occurs. The project will determine how Notchl regulates the genomes of T-ALL cells, both in general and with specific genes and co-factors. In doing so, this project will uncover molecular mechanisms that will lead to new diagnostic and therapeutic opportunities in T-ALL and other Notch-related cancers.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Program Projects (P01)
Project #
5P01CA119070-08
Application #
8701032
Study Section
Special Emphasis Panel (ZCA1)
Project Start
Project End
Budget Start
2014-07-01
Budget End
2015-06-30
Support Year
8
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
City
Boston
State
MA
Country
United States
Zip Code
02115
Stein, Sarah J; Mack, Ethan A; Rome, Kelly S et al. (2016) Trib2 Suppresses Tumor Initiation in Notch-Driven T-ALL. PLoS One 11:e0155408
Bernasconi-Elias, P; Hu, T; Jenkins, D et al. (2016) Characterization of activating mutations of NOTCH3 in T-cell acute lymphoblastic leukemia and anti-leukemic activity of NOTCH3 inhibitory antibodies. Oncogene 35:6077-6086
Chiang, Mark Y; Wang, Qing; Gormley, Anna C et al. (2016) High selective pressure for Notch1 mutations that induce Myc in T-cell acute lymphoblastic leukemia. Blood 128:2229-2240
Guo, Bingqian; McMillan, Brian J; Blacklow, Stephen C (2016) Structure and function of the Mind bomb E3 ligase in the context of Notch signal transduction. Curr Opin Struct Biol 41:38-45
McMillan, Brian J; Tibbe, Christine; Jeon, Hyesung et al. (2016) Electrostatic Interactions between Elongated Monomers Drive Filamentation of Drosophila Shrub, a Metazoan ESCRT-III Protein. Cell Rep 16:1211-7
Xu, Xiang; Choi, Sung Hee; Hu, Tiancen et al. (2015) Insights into Autoregulation of Notch3 from Structural and Functional Studies of Its Negative Regulatory Region. Structure 23:1227-35
McMillan, Brian J; Schnute, Björn; Ohlenhard, Nadja et al. (2015) A tail of two sites: a bipartite mechanism for recognition of notch ligands by mind bomb E3 ligases. Mol Cell 57:912-24
Wang, Hongfang; Zang, Chongzhi; Liu, X Shirley et al. (2015) The role of Notch receptors in transcriptional regulation. J Cell Physiol 230:982-8
Gordon, Wendy R; Zimmerman, Brandon; He, Li et al. (2015) Mechanical Allostery: Evidence for a Force Requirement in the Proteolytic Activation of Notch. Dev Cell 33:729-36
Wang, Hongfang; Zang, Chongzhi; Taing, Len et al. (2014) NOTCH1-RBPJ complexes drive target gene expression through dynamic interactions with superenhancers. Proc Natl Acad Sci U S A 111:705-10

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